GI38 
6 


• 


THE  LIBRARY 

OF 

THE  UNIVERSITY 
OF  CALIFORNIA 

PRESENTED  BY 

PROF.  CHARLES  A.  KOFOID  AND 
MRS.  PRUDENCE  W.  KOFOID 


GASTRIC  DERANGEMENTS. 


HORSFORD'S  ACID  PHOSPHATE. 


Unlike  all  other  forms  of  phosphorus  in  combination,  such  as  dilute 
phosphoric  acid,  glacial  phosphoric  acid,  neutral  phosphate  of  lime,  hypo- 
phosphites,  etc.,  the  phosphates  in  this  product  are  in  solution,  and  readily 
assimilated  by  the  system,  and  it  not  only  causes  no  trouble  with  the 
digestive  organs,  but  promotes  in  a  marked  degree  their  healthful  action. 

In  certain  forms  of  dyspepsia  it  acts  as  a  specific. 

Dr.  T.  G.  COMSTOCK,  of  the  Good  Samaritan  Hospital,  St.  Louis, 
says:  "  For  some  years  we  have  used  it  in  a  variety  of  derangements 
characterized  by  debility,  as  also  in  chronic  gastric  ailments.  It  is  ap- 
proved of,  unanimously,  by  the  medical  staff  of  this  Hospital." 


Send  for  descriptive  circular.     Physicians  who  wish  to  test  it  will  be 
furnished  a  bottle  on  application,  without  expense  except  express  charges. 
Prepared  under  the  direction  of  Prof.  E.  N.  HORSFORD,  by  the 

RUMFORD  CHEMICAL  WORKS,  Providence,  R.  L 


Beware  of  Substitutes  and  Imitations. 


UBKf 

COUFGf  'M 


ThjE1 

Bacterial  Poisons. 


BY 

DR.   N.    GAMALEIA. 


TRANSLATED  BY  E.  P.  HURD,  M.D., 

Member  of  the  Massachusetts  Medical  Society  and  of  the  Climatological 

Society ;  one  of  the  Physicians  to  the  Newburyport  {Mass^ 

Hospital. 


GEORGE  S.  DAVIS, 

DETROIT,  MICH. 


80 


Copyrighted  by 

GEORGE  ?.  DAVIS. 

1898. 


TABLE  OF  CONTENTS. 


FIRST     PART. 
CHAPTER  I. 

EXPERIMENTAL  STUDY  ON  THE  PUTRID  POISON. 

PAGE 

Experiments  with  the  Toxic  Effects  Produced  by  Sub- 
stances Undergoing  Decomposition — Seybert,  Gas- 
pard,  Stich  Establish  the  Symptoms  and  the  Lesions 
of  Experimental  Septicaemia— Panum  Gives  the 
Proof  that  it  is  Produced  by  a  Chemical  Poison — 
Bergmann  and  Schmiedeberg  Determine  this  Poi- 
son to  be  an  Alkaloid:  Sepsine I 

CHAPTER  II. 

THE    MICROBIAL   ETIOLOGY  OF   PUTREFACTION   AND 
OF  THE  INFECTIONS. 

Contest  between  the  Chemical  and  the  Vitalist  Doctrines 
— The  Contradictions  and  the  Results  Obtained  by 
the  Chemists — Their  Explanation  Insufficient  by 
Reason  of  the  Absence  of  the  Notion  of  Specificity — 
Definitive  Victory  of  the  Vitalist  Doctrine — The  In- 
fectious Diseases  are  Admitted  to  be  Caused  by 
Specific  Fermentations 12 

CHAPTER  III. 
THE  DISCOVERY  OF  PTOMAINES. 

Selmi  Draws  Attention  to  the  Ptomaines  Obtained  from 
Cadavers — Leucomaines  and  Ptomaines  of  Gautier 
— The  Researches  of  Brieger  on  Ptomaines 20 


VI 

PAGE 
CHAPTER  IV. 

INFECTION  is  AN  INTOXICATION  BY  THE  MICROBIAN 
POISON. 

Bacteriology  Aims  to  Explain  Infection  by  the  Invasion 
of  the  Animal  Economy  by  Microbes — Three  Dis- 
eases, Diphtheria,  Cholera,  Tetanus,  where  this  Ex- 
planation is  Not  Possible — Discovery  of  Chemical 
Vaccination — Discovery  of  the  Different  Chemical 
Poisons  of  Bacteria  which  May  Produce  All  the 
Symptoms  of  the  Disease  without  the  Intervention 
of  Microbes — Actual  Point  of  View:  The  Infectious 
Disease  is  an  Intoxication  by  the  Poison  of  the 
Pathogenic  Microbe 30 


SECOND     PART. 
CHAPTER  V. 

THE  CHEMICAL  NATURE  OF  THE  BACTERIAL  POISONS. 

Different  Conceptions  of  the  Chemical  Nature  of  the 
Bacterial  Poisons.  First  Stage:  The  Ptomaines — 
Differences  between  the  Ptomaines  of  Selmi,  of 
Gautier,  and  of  Brieger — The  Peptotoxine  of  Brieger 
— Researches  of  Salkowsky,  of  Bouveret  and  Devic 
— Researches  of  Bassi  on  Typhotoxine — Works  of 
Baumann — The  Ptomaines  are  Perhaps  Artificial 
Products — Opinion  of  S.  Martin.  Second  Stage: 
The  Diastases — Researches  of  Roux  and  Yersin 
— Criticism  of  the  Notion  of  Diastases.  Third 
Stage:  The  Toxalbumins — Work  of  Brieger  and 
Frankel 39 


VII 

PAGE 
CHAPTER  VI. 

THE  CHEMICAL  NATURE  OF  THE  BACTERIAL  POISONS 
(CONTINUED). 

Criticism  of  the  Work  of  Brieger  and  Frankel — Re- 
searches of  Proskauer  and  Wassermann — The  Mi- 
crobe Poisons  have  Not  yet  been  Prepared  in  a  State 
of  Purity — Similar  Non-Microbian  Poisons — The 
Work  of  Stillmarck — Demonstration  of  the  Albu- 
minoid Nature  of  the  Diphtheritic  Poison — Toxi- 
cological  Differentiation  of  the  Bacterial  Poisons — 
Natural  and  Artificial  Poisons:  Their  Characters — 
The  Chemical  Vaccines — The  Antitoxines  or  Cura- 
tive Substances — Hypothesis  as  to  the  Chemical 
Nature  of  the  Poisons — The  Nucleo-albumins  and  the 
Nucleines — The  Consequences  of  this  Hypothesis.. .  52 

CHAPTER  VII. 
THE  ORIGIN  OF  THE  MICROBIAN  POISONS. 

Preconceived  Idea  that  the  Poisons  Can  Only  Come 
from  the  Decomposition  of  Albuminoid  Bodies — Ex- 
periments Already  Old  which  Overthrow  this  Idea — 
Researches  of  Polotebnoff,  Popoff,  Bergmann,  Schul- 
ler — Numerous  Researches  Made  at  the  Laboratory 
of  Pachoutine — Recent  Labor  of  Guinochet — The 
Microbian  Poisons  are  Not  Products  of  Decompo- 
sition, but  the  Result  of  Synthesis — Are  They  Se- 
cretions?— Arguments  in  Favor  of  this  Idea — Argu- 
ments which  Combat  it — Thje  Poisons  are  Intimately 
Linked  to  the  Bodies  of  the  Bacteria — The  Experi- 
ments of  Cantani  and  of  the  Author — The  Re- 
searches of  Buchner  on  the  Proteins— Criticism  of 
these  Researches — All  the  Microbe  Poisons  Come 


VIII 

PAGE 

from  the  Bodies  of  Bacteria — Utility  of  this  Source 
for  the  Microbes 65. 

CHAPTER  VIII. 

ACTION  OF  THE  BACTERIAL  POISONS  ON  THE  ANIMAL 
ORGANISM — ACCUSTOMANCE  AND  IMMUNITY. 

Insufficiency  of  our  Knowledge  of  the  Mode  of  Action  of 
the  Bacterial  Poisons  on  the  Animal  Organism — 
Local  Action  of  the  Modified  Poisons — Selective  Ac- 
tion of  Tuberculin  and  Malleine — Destruction  of 
Poisons  in  the  Bodies  of  Refractory  Animals — 
Explanations  of  the  Different  Contradictions — 
Resume  and  Conclusions  on  the  General  Toxicity  of 
Microbes 78 


THIRD     PART. 

CHAPTER  IX. 
THE  POISONS  OF  TETANUS. 

Researches  of  Brieger,  Kitasato  and  Weyl,  Knud  Faber, 
Tizzoni  and  Cattani,  Brieger  and  Frankel,  Vaillard 
and  Vincent  and  Kitasato,  on  the  Properties  of  the 
Tetanus  Poison — The  Investigations  of  Bruschetini 
and  of  Camara  Pestana  on  its  Diffusion  in  the  Ani- 
mal Body — Vaccination  against  Tetanus:  Behring 
and  Kitasato — The  New  Notions  Contributed  by 
these  Authors 88 

CHAPTER  X. 

THE  POISONS  OF  TETANUS  (CONTINUED). 
Vaccination   against   Tetanus   is   Obtained    by  Tizzoni, 


IX 

PAGE 
Cattani,  and  Vaillard — The  Researches  of  Kitasato; 

of  Behring;  of  Brieger,  Kitasato,  and  Wassermann 
— Immunization — The  Labors  of  Tizzoni,  Cattani, 
and  Vaillard — The  Researches  of  Ehrlich;  of  Brieger 
and  Ehrlich;  of  Brieger  and  Frank — Application  of 
the  Method  of  Behring  and  Kitasato  to  the  Treat- 
ment of  Tetanus  in  Man 97 

CHAPTER  XI. 

•» 
THE  POISONS  OF  DIPHTHERIA. 

The  Researches  of  Roux  and  Yersin;  of  Loffler — C. 
Frankel  Succeeds  in  Vaccinating  Animals  against 
Diphtheria — The  Researches  of  Behring  and  His 
Discovery  of  Immunization — The  Difficulties  Not 
Yet  Surmounted  of  Diphtheritic  Vaccination 106 

CHAPTER  XII. 

THE   POISONS    OF   CHOLERA   AND   OF    THE    AVICIDE 
VIBRIO. 

The  Researches  of  the  Author  on  the  Poisons  of  Cholera 
and  of  the  Avicide  Vibrio — The  Researches  of  Her- 
nandez and  Brtthl,  of  Niessen  and  Behring,  and  of 
Zasslein 1 14 

CHAPTER  XIII. 
THE  POISONS  OF  TUBERCULOSIS. 

The  Researches  of  Koch,  of  Maffuci,  of  Prudden  and 
Hodenpyl,  of  Straus  and  the  Author,  of  Grancher 
and  Ledoux-Lebard — The  Tuberculin  of  Koch,  and 
the  Researches  which  it  has  Provoked — The  Toxo- 
mucin  of  Weyl 120 


X 

PAGE 
CHAPTER  XIV. 

THE  POISONS  OF  CHARBON  AND  OF  GLANDERS. 

Insufficiency  of  our  Toxicological  Knowledge  respecting 
Charbon  and  Glanders — Different  Writings  on  the 
Subject 127 

CHAPTER  XV. 

PRACTICAL    RESULTS    OBTAINED    BY   THE   STUDY  OF 
MICROBIAN  TOXICOLOGY. 

Rapid  Indications  respecting  the  Other  Bacterial  Poisons 

— Immunization  as  a  Result  of  Toxicological  Studies.   133 


PREFACE. 


The  Bacterial  Poisons  are  here  for  the  first  time  made 
the  subject  of  a  connected  treatise.  The  genera]  plan  of  this 
work  is  as  follows: 

When  bacteriology,  extending  its  domain  beyond  certain 
etiological  data  concerning  infection,  sought  to  penetrate  the 
pathogeny  thereof,  there  arose  a  necessity  of  undertaking 
the  study  of  chemical  poisons  produced  by  the  microbes.  This 
study  first  imposed  itself  in  diseases  such  as  cholera,  diphthe- 
ria, and  tetanus,  where  the  pathogenic  agent  is  found  lodged 
in  a  limited  part  of  the  organism  and  yet  causes  a  general 
affection.  In  these  cases,  the  pathogenic  action  of  the  bac- 
teria can  be  explained  only  by  the  systemic  poisoning  pro- 
duced by  the  specific  products  of  these  bacteria.  But  the 
same  explanation  has  been  found  to  hold  good  in  diseases  of 
another  type,  where,  as  in  tuberculosis,  the  lesion  produced 
by  the  pathogenic  agent  is  strictly  circumscribed  to  the  spot 
where  this  agent  vegetates;  here  also  we  have  proved  that  the 
lesion  does  not  come  from  the  microbe  as  a  living  organism, 
but  from  its  chemical  poisons — the  lesion  and  the  disease 
may  be  reproduced  by  the  dead  microbe,  which  can,  of  course, 
act  only  by  the  toxic  substances  which  it  contains. 

The  study  of  bacterial  poisons  concerns  not  only  the 
pathogeny  of  infectious  diseases;  it  finds  also  practical  appli- 
cations for  their  prophylaxis  and  treatment.  It  has  been 
demonstrated  that  a  sure  and  inoffensive  vaccination  may  be 
obtained  without  the  help  of  living  microbes,  and  that  the 
refractory  state  may  be  conferred  simply  and  solely  by  the 
soluble  products  of  micro-organisms.  Lastly,  a  new  method 
has  been  found — immunization — which  has  direct  applica- 


XII 

tions  in  therapeutics,  being  capable  not  only  of  preventing 
disease,  but  of  curing  it  when  once  declared.  Here  the  role 
of  living  bacteria  is  even  less  than  in  chemical  vaccination. 
The  cure  of  infectious  diseases  is  obtained  with  the  serum  of 
vaccinated  animals. 

The  theory  of  immunity  has  also  profited  by  the  knowl- 
edge of  the  poisons  of  bacteria;  it  has  been  shown  that  the 
immunity  of  animals  against  infectious  diseases  is  in  close 
relation  with  their  resistance  to  microbial  poisons;  the  latter 
is  subordinated  in  its  turn  to  the  elaboration  by  these  ani- 
mals of  special  substances — antitoxines — which  have  the 
property  of  neutralizing  the  action  of  microbial  toxines. 

From  this  aggregate  of  facts,  a  new  science  has  sprung 
up:  the  science  of  Microbial  Poisons,  which  is  based  at  once 
on  bacteriology,  on  biological  chemistry,  and  on  general 
physiology. 

From  bacteriology  it  borrows  its  data  about  microbes  as 
producers  of  poisons;  but  the  important  results  obtained  with 
the  analogous  poisons  of  different  origin,  such  as  abrine, 
ricine,  robine,  necessitate  the  extension  of  the  domain  of 
microbial  toxicology  beyond  microbism. 

To  biological  chemistry  belong  the  methods  of  prepara- 
tion and  of  isolation  of  bacterial  poisons;  from  it  we  must 
demand  information  as  to  their  nature  and  their  production. 

Lastly,  the  study  of  their  mode  of  action  on  the  animal 
•organism,  of  their  diffusion,  and  of  their  destruction  in  the 
organism,  belongs  to  the  domain  of  general  physiology. 

Thus,  microbial  toxicology  touches  at  once  on  bacteriol- 
ogy, on  biological  chemistry,  and  on  general  physiology;  it 
is  the  science  of  bacterial  poisons,  and  has  for  its  object  the 
-study  of  their  chemical  nature,  of  their  mode  of  production, 
.and  of  their  action  on  animals. 

This  work  is  divided  into  three  parts: 

The   first  part — History  of  Microbial  Toxicology  —  de- 


XIII 

scribes  the  evolution  which  the  study  oi  systemic  poisoning 
in  the  infections  has  undergone. 

The  second  part — General  Toxicology — treats  of  our 
actual  information  respecting  the  chemical  nature  of  the 
microbial  poisons,  their  production,  and  their  destiny  in  the 
animal  body. 

The  third  part — Special  Toxicology — will  be  the  exposi- 
tion of  the  data  acquired  concerning  the  toxines  of  different 
•diseases.  N.  G. 

Paris,  May,  1892. 


THE  BAOTEKIAL  POISONS. 

First  Part:  History  of  the  Development  of 

Our  Knowledge  Respecting  the 

Bacterial  Poisons. 


CHAPTER  I. 

EXPERIMENTAL  STUDY  ON  THE   PUTRID 
POISON. 


SUMMARY. — Experiments  with  the  Toxic  Effects  Pro- 
duced by  Substances  Undergoing  Decomposition — 
Seybert,  Gaspard,  Stick  Establish  the  Symptoms  and 
the  Lesions  of  Experimental  Septicczmia — Panum 
Gives  the  Proof  that  it  is  Produced  by  a  Chemical 
Poison — Bergmann  and  Schmiedeberg  Determine 
This  Poison  to  be  an  Alkaloid:  Sepsine. 

The  first  microbial  poisons  studied  experimentally 
were  products  of  putrefaction.  Chemists  were  led  to 
this  study  by  the  important  etiological  role  which 
putrefaction  plays  in  ancient  medicine.  It  was  be- 
lieved that  the  emanations  from  decomposing  matters 
might  produce  typhoid  and  malarial  fevers.  The 

I  000 


putrefaction  of  wounds  gave  rise  to  the  complications 
so  formidable  and  so  frequent,  of  septicaemia  and 
pyaemia.  A  "spontaneous"  tendency  of  the  living 
organism  to  putridity  characterized  certain  wide- 
spread diseases — such,  for  instance,  as  scorbutus. 
The  diseases  of  putrid  origin  thus  constituted  an 
important  group  of  affections  which  corresponded 
nearly  to  our  infectious  diseases  of  to-day. 

Despite  this  importance  which,  from  an  early 
antiquity,  medicine  accorded  to  putrefaction,  the 
experimental  study  of  the  latter  is  relatively  recent. 

Towards  the  end  of  the  last  century,  §£gtejj|^£.-  jr» 
perimented  with  blood  serum,  pus,  and  putrid  meat. 
He  introduced  these  matters  into  the  general  circula- 
tion of  the  dog.  He  noticed  that  the  toxic  effects 
varied  proportionately  to  the  quantities  of  liquid  in- 
jected. Thus,  for  instance,  20  cubic  centirneters  of 
putrid  serum  killed  a  dog  Jn_the_j^ace7flL~several 
hours,  with  vomitings,  convulsions,  and  progressive  en- 
feeblement;  3  grammes  caused  deathjrrtwo  da^s,  with 
dysenteric  pnenomefrar^  doses  below  two  grammes 
produced  only  a  temporary  diarrhoea,  followed  by 
a  complete  restoration  of  the  animal.  Seybert  also 
studied  the  effects  of  the  same  putrescent  sub- 
stances when  they  were  introduced  by  mouth  into 
the  stomach.  In  these  cases  the  toxic  effects  were 
nil.  Moreover,  in  opening  the  stomach  of  the  dog 
three  hours  after  a  meal  of  putrid  meat,  Seybert  no 
longer  found  in  the  mass  undergoing  digestion  the 


——      •?          ..-. 

putrid  aspect  and  odor.  Seybert  cites  in  this  connec- 
tion the  researches  of  Spalanzani,  who  was  absolutely 
convinced  of  the  innocuousness  of  putrid  foods  after 
having  fed  dogs,  cats,  and  birds  with  them. 

Gaspard,  physician  of  Saint  Etienne,  pursued 
some  experimental  researches  on  the  putrid  poison. 
His  experiments  established  several  important  points 
respecting  the  effects  of  putrid  matters. 

y  Gaspard  showed  that  (fafiflmp™1^  blood,  pus, 
and  meat  produce  in  animals  a  rapid  and  mortal 
intoxication.  On  the  contrary,  fresh  organic  liquids 
such  as  saliva,  urine,  and  sperm,  when  injected  for 
control,  are  found  to  be  non-toxic.  Gaspard  showed, 
moreover,  that  the  putrid  infusions  of  plants  also 
poison  animals. 

The  symptoms  of  this  poisoning  were  very  con- 
stant and  typical,  being  independent  of  the  source  of 
the  putrid  substances,  whether  vegetable  or  animal, 
and  of  the  place  of  the  injection.  This  intoxication 
(septic  or  putrid  infection)  was  characterized  by 
tremblings  and  convulsions,  vomitings,  and  diarrhoea, 
the  latter  often  sanguinolent.  The  animals  died  in 
dyspnoea,  cyanosis,  and  complete  prostration. 

At  the  autopsy,  the  pathognomonic  lesion  of  the 
putrid  infection  consisted  in  a  haemorrhagic  enteritis. 

Gaspard  concluded  from  his  experiments  that  a 
special  poison,  the  putrid  poison,  is  formed  by  putre- 
faction. This  poison  produces  the  same  disease  and 
the  same  lesions  in  all  the  animal  species. 


—  4  — 

In  order  to  determine  the  nature  of  the  putrid 
poison,  Gaspard  experimented  with  the  different 
gases  formed  by  putrefaction.  He  found  that  car- 
bonic acid  and  sulphuretted  hydrogen  are  not 


that  ammonia,  on  the  contrary,  is  toxic,  but  that  it 
cannot  produce  the  symptoms  of  putrid  infection^ 
Hence  was  overthrown  the  preconceived  idea  of  the 
ancients,  who  believed  the  principal  danger  to  be  in 
the  putrid  gases  of  decomposition.  The  researches 
of  Gaspard  established  subsequently  the  formation, 
in  the  course  of  putrefaction,  of  a  special  and  very 
active  poison.* 

Other  experimenters  followed  in  the  way  marked 
out  by  Gaspard.  Magendie,  Leuret,  Dupuis,  Darcet, 
and  Sedillot  confirmed  on  all  the  principal  points  the 
fundamental  reseaches  of  Gaspard.  They  discovered 
his  putrid  poison,  without,  however,  determining  its- 
chemical  nature. 

Virchow  found  that  the  putrid  poison  does  not 
act,  like  the  other  chemical  poisons,  in  proportion  to- 
rts quantity.  Its  activity  depends  rather  on  the  de- 
gree of  decomposition  of  the  substances  whence  it  is 
derived.  From  this  point  of  view,  Virchow  ranks  it 
among  the  ferments. 

Stich  has  made  very  careful  and  interesting  re- 
'Searches  on  putrid  matters.  He  injected  putrefied 
albuminoid  products,  after  repeated  filtration  through- 

*  Gaspard:  Physiological  Memoir  on  the  Purulent  and. 
Putrid  Maladies  (Journal  de  Physiologic,  1822  and  1824). 


T      r*        

paper,  into  the  veins  of  dogs,  of  hares,  and  of  birds. 
The  latter  showed  themselves  the  most  susceptible  to 
poisoning.  The  typical  lesion  consisted  in  a  hyper- 
semia  of  the  intestines. 

But  especially  interesting  are  the  researches  of 
Stich  on  the  toxicity  of  faecal  matters.  The  aqueous 
extract  of  the  excrement  of  a  dog,  injected  into  the 
veins  of  this  animal,  was  found  to  be  very  toxic;  but 
the  same  extract  introduced  into  the  stomach  or  rec- 
tum of  dogs  was  completely  inoffensive. 

Stich  carried  this  study  farther.  Introducing 
the  excrement  of  one  animal  into  the  stomach  or 
rectum  of  an  animal  of  another  species,  he  found  that 
poisoning  was  the  result.  It  follows  that  immunity 
exists  for  animals  only  in  relation  to  their  own  in- 
testinal contents.  Stich  asks:  What  are  the  causes  of 
this  absence  of  auto-intoxication?*  He  leaves  the 
question  unsolved. 

The  most  important  researches  on  the  putrid 
poison  belong  indisputably  to  Panum. 

J^i^rr^asks  if  the  accidents  of  putrid  infection 
are  really  due  to  a  chemical  substance  or  to  the  action 
of  bacteria  multiplying  in  matters  undergoing  putre- 
faction? To  solve  this  question,  he  instituted  numer- 
ous important  researches  on  the  putrid  poison,  and 
confirmed  the  results  obtained  by  Gaspard. 
duced  a  typical  intofijfiflt^"  *T  **"»- '"-f^*^  nf 
fied  meat.  This  intoxication  was  characterized  by 

*  Stich,  Charite  Annalen,  1853. 


K 


prostration,  vomiting  and  diarrhoea,  and  rapid  death. 
The  autopsy  showed  a  more  or  less  pronounced 
gastro-intestinal  catarrh. 

Having  reproduced  this  putrid  infection  of  Gas- 
pard,  Panum  then  demonstrated  that  it  was  due  to 
the  action  of  a  chemical  poison  independent  of  the 

^pjpO^g^^gj^ggjBjpgj^^^gjgfcMefMVHMUMMMPMMMI  .         ..*«*  * 

agency  of  bacteria.  The  putrid  liquids,  though  per- 
fectly clear,  and  freed  from  every  foreign  germ  by  re- 
peated filtration  through  paper,  none  the  less  retained 
their  toxic  properties.  Nor  was  the  putrid  poison  de- 
stroyed by  ebullition  for  eleven  hours,  which  ought 
surely  to  kill  all  living  germs.  Having  thus  settled 
the  fundamental  question  as  to  the  inorganic  charac- 
ter of  the  putrid  poison,  Panum  studied  its  chemical 
properties.  He  found  that  this  poison  was  not  vola- 
tile; for  the  distilled  products  of  the  putrefied  matters,, 
while  having  the  fetid  odor,  are  not  at  all  toxic. 
The  putrid  poison  was  fixed,  and  the  dry  residue ^of 
the  evaporation  of  the  putrefied  matters  retained  all 
their  toxic  properties..  This  dried  residue  did  not 
give  up  to  alcohol  its  chief  toxic  principle.  The  alco- 
holic extract  had  a  different  physiological  action  from 
the  putrid  matters;  it  produced  a  manifest  narcosis,, 
which  Panum  compared  to  that  produced  by  the 
alkaloids  of  opium.  The  veritable  putrid  poison 
capable  of  causing  the  typical  poisoning  was,  on  the 
contrary,  soluble  in  water,  although  it  adhered  to  the 
precipitates  of  the  albuminoid  substances  coagulable 
by  heat;  it  was  also  partially  retained  by  the  filters. 


L  t        7     __ __ 

Panum  showed  also  that  his  poison  had  nothing  in 
common  with  the  well  known  substances  produced  by 
putrefaction,  such  as  leucin  and  tyrosin.  He  stopped 
here  in  his  chemical  investigation,  and  did  not  further 
indicate  the  nature  of  his  putrid  poison.  It  was  suffi- 
cient for  him  to  have  given  the  irrefutable  proof  that 
it  was  not  organized. 

The  work  of  Panum  produced  an  enormous  im- 
pression upon  the  scientific  world.  Hidden  by  its 
author  in  an  unknown  Danish  publication,  the  memoir 
of  Panum  was  found  there  by  a  German  writer, 
who  published  an  analysis  of  it  in  Schmitt's  Jahr- 
biicher  for  1859.  In  this  extensively  circulated  pub- 
lication the  results  of  Panum's  researches  became 
known  to  the  whole  scientific  world.  A  great  num- 
ber of  experimenters  in  Germany  and,  especially,  in 
Russia,  occupied  themselves  with  the  verification  of 
these  results  and  the  chemical  study  of  the  putrid 
poison. 

The  University  of  Munich  then  offered  a  prize 
for  the  best  essay  on  "  Putrid  Infection  and  Its 
Causes."  Two  memoirs  were  deemed  worthy  of  a 
prize:  those  of  Henner  and  Schweninger.  These 
writers  confirmed  the  fundamental  result  of  Panum, 
that  the  putrid  poison  is  of  chemical  nature,  and  that 
the  action  of  microbes  is  nil  in  putrid  infection.  As 
to  the  nature  of  the  poison,  they  favor  Virchow's 
view  that  it  should  be  ranked  among  the  ferments,  for 
it  acts  in  infinitesimal  quantity,  has  a  period  of  incu- 


bation,  and  always  produces  the  same  typical  affec- 
tion. 

Since  the  same  year,  1866,  there  have  appeared 
at  Dorpat  numerous  theses  devoted  to  the  chemi- 
cal study  of  the  putrid  poison.  We  may  cite  the 
memoirs  of  Raison,  Frese,  Weidenbaum,  Schmitz, 
Petersen,  A.  Schmidt,  and  Brehm.  These  theses  also 
confirm  Panum's  position.  Raison  shows  that  the 
putrid  poison  is  not  retained  by  filtration  through 
charcoal.  The  minimum  dose  of  0.0036  gramme  of 
the  filtered  liquid  killed  a  horse  by  intravenous  in- 
jection. He  found  also  that  the  putrid  poison,  evap- 
orated to  dryness,  could  be  subjected  for  several 
hours  to  a  heat  of  130°  C.  Weidenbaum  has  seen 
that  the  putrid  poison  supports  discontinuous  ebulli- 
tion, repeated  several  times.  These  two  writers,  and 
the  other  pupils  of  the  school  of  Dorpat  whom  we 
have  mentioned,  have  also  seen  the  putrid  poison 
support  without  alteration  the  different  chemical 
manipulations,  such  as  the  action  of  sulphuric  and 
hydrochloric  acids,  precipitation  by  acetate  of  lead 
and  nitrate  of  mercury,  etc.  Lastly,  they  even  suc- 
ceeded in  obtaining  the  putrid  poison  in  a  state  of 
purity,  and  of  determining  its  chemical  nature.  It  is 
the  celebrated  sepsine  found  in  1868  by  Bergmann 
and  Schmeideberg.  This  important  result  was  ac- 
complished by  a  complicated  method  based  on  the 
employment  of  corrosive  sublimate. 

In  1866,  Bergmann  published  the  results  of  his 


—  9  — 

researches  made  with  Schmiedeberg  on  the  chemical 
properties  of  the  poison  contained  in  putrefied  yeast. 
They  found  that  this  poison  supports  for  eight  hours 
digestion  in  boiling  alcohol.  The  alcoholic  extract 
of  the  putrefied  yeast  is  found,  after  evaporation,  to  be 
powerfully  toxic.  To  separate  the  active  principle, 
Bergmann  and  Schmiedeberg  precipitated  it  in  alco- 
holic solution  by  corrosive  sublimate.  The  mercurial 
precipitate  was  washed  in  alcohol,  dissolved  in  water, 
and  decomposed  by  hydrogen  sulphide.  The  liquid, 
freed  from  sulphide  of  mercury  by  filtration,  was 
heated  to  get  rid  of  the  excess  of  hydrogen  sulphide, 
and  treated  with  carbonate  of  silver,  which,  by  form- 
ing an  insoluble  chloride  of  silver,  removed  free 
hydrochloric  acid  from  the  solution.  The  filtered 
liquid  obtained  after  this  last  operation  was  almost 
colorless  and  perfectly  clear.  It  had  the  specific 
toxicity  of  the  putrid  poison. 

In  1868  these  experimenters  (Bergmann  and 
Schmiedeberg)  obtained,  in  crystalline  form,  the  salt 
of  this  poison  with  sulphuric  acid,  and  called  'it  the 
sulphate  of  sepsine.  This  salt  was  proved  to  be 
eminently  toxic.  Injected  in  the  dose  of  one  centi- 
gramme in  the  vein  of  the  dog,  it  provoked  imme- 
diately vomiting  and  sanguinolent  diarrhoea,  and  at 
the  autopsy  hsemorrhagic  ecchymoses  were  found  in 
the  stomach  and  intestines. 

The  publication  of  Bergmann  and  Schmiedeberg's 
discoveries  naturally  produced  a  profound  impres- 


IO 


sion.  For  a  time,  chemists  believed  they  had  at  last 
got  hold  of  the  famous  putrid  poison  whose  existence 
had  been  demonstrated  by  Gaspard,  whose  purely 
chemical  nature  had  been  proved  by  Panum,  and  which 
had  finally  been  obtained  in  a  crystalline  form  by  Berg- 
mann  and  Schmiedeberg.  This  sepsine  would  alone 
henceforth  explain  all  the  various  accidents  of  medi- 
cal and  surgical  septicaemia.  It  was  believed  that  the 
way  was  finally  opened  for  the  easy  interpretation  of 
all  the  infections.  Chemists  went  to  work  to  seek 
out  and  to  study  this  sepsine.  Petersen  and  A. 
Schmidt  found  it  in  putrid  blood,  and  thus  confirmed 
the  data  of  Bergmann  and  Schmiedeberg. 

Nevertheless,  the  most  competent  authorities 
soon  admitted  the  impossibility  of  ascribing  all  the 
effects  of  the  putrid  poison  to  sepsine  alone.  First,  it 
is  not  found  in  all  putrefied  substances;  thus,  for  in- 
stance, Fischer  could  not  find  it  in  putrid  pus.  Berg- 
mann himself,  in  applying  his  method  of  extraction 
to  different  putrid  matters,  has  isolated  substances 
which  had  nothing  in  common  with  sepsine.  Other 
toxic  substances  were  extracted  from  the  putrid 
matters. 

Zuelzer  and  Sonnenschein  have  found  another 
septic  alkaloid  in  the  infusions  of  putrid  meat.  This 
body  has  all  the  reactions  of  the  vegetal  alkaloids, 
such  as  atropine  and  hyoscyamine.  It  possesses  also 
the  physiological  action  of  the  latter.  Injected  under 
the  skin  of  animals,  it  provokes  dilatation  of  the 


pupils,  relaxation  of  the  intestines,  and  exaltation  of 
the  heart's  action. 

The  conclusion  was  inevitable  that  the  poisons 
produced  by  putrefaction  were  numerous  and  differ- 
ent, according  to  conditions  necessary  to  determine. 
In  the  following  chapter  we  shall  study  the  researches 
which  have  been  undertaken  to  elucidate  the  rationale 
of  putrefaction  and  of  septicaemia. 

Only,  we  shall  see  in  studying  the  different  pos- 
sible causes  which  influence  the  production  of  the 
putrid  poison,  that  experimenters  had  overlooked  the 
principal  one,  viz.,  the  intervention  of  microbes.  Be- 
tween the  Chemical  School  and  the  Vitalists,  there 
was  destined  to  be  a  memorable  struggle,  which 
ended  in  the  victory  of  the  latter — a  victory  which 
completely  cast  into  the  shade  the  results  acquired  by 
the  chemical  study  of  the  putrid  poison. 


CHAPTER  II. 

THE  MICROBIAL  ETIOLOGY  OF  PUTREFACTION 
AND  OF  THE  INFECTIONS. 


SUMMARY.  —  Contest  between  the  Chemical  and  the 
Vitalist  Doctrines — The  Contradictions  and  the 
Results  Obtained  by  the  Chemists — Their  Explana- 
tion Insufficient  by  Reason  of  the  Absence  of  the 
Notion  of  Specificity — Definitive  Victory  of  the 
Vitalist  Doctrine — The  Infectious  Diseases  are  Ad- 
mitted to  be  Caused  by  Specific  Fermentations. 

From  antiquity,  two  contrary  interpretations  of 
the  etiology  of  putrid  and  infectious  diseases  have 
prevailed. 

The  Chemical  Doctrine  set  forth  in  the  previous 
chapter  is,  that  matters  in  the  process  of  decomposi- 
tion constitute  poisons  sufficient  to  account  for  all  the 
symptoms  of  the  infections,  and  that  the  microbes 
found  in  these  putrid  matters  are  only  inconstant  and 
inoffensive  accompaniments.  The  other,  or  Germ 
Theory,  affirms  that  the  microbes  are  really  the  sole 
agents  of  putrefaction  and  of  infectious  diseases,  and 
that  just  as  the  inanimate  matters  do  not  alter  and  fer- 
ment without  the  intervention  of  microbes,  so  the 
presence  of  the  latter  is  indispensable  for  the  produc- 


—  13  — 

tion  of  the  infectious  disease.  We  shall  now  see  how 
this  Vitalist  doctrine  has  definitively  supplanted  the 
Chemical. 

The  chemical  and  toxicological  researches  on  the 
putrid  poisons  did  not  realize  the  fundamental  condi- 
tion of  all  scientific  work;  they  did  not  give  results 
that  were  always  constant  and  uniform.  Chemical 
analysis  revealed  different  properties  in  the  active 
principles  isolated.  Experimentation  in  the  hands  of 
different  chemists  did  not  produce  the  same  symptoms 
and  the  same  lesions  with  the  poisons  obtained  from 
putrid  substances.  This  inconstancy  of  results  was 
soon  noticed  by  investigators.  Thus,  for  instance, 
one  of  them  *  expresses  himself  in  this  way: 

"  There  is  not  agreement  as  to  the  pathology  of 
the  putrid  infection  or  as  to  its  nature.  Thus,  cer- 
tain chemists  (Henner,  Schmidt,  Petersen)  affirm  that 
the  convulsions  and  tetanic  attacks  are  important 
characters  of  putrid  infection;  others,  on  the  contrary 
(Dupuis,  Sedillot,  Stick,  Raison,  and  Schmitz),  have 
never  or  but  rarely  seen  them.  Even  the  constant 
existence  of  vomiting  and  diarrhoea,  supposed  to  be 
pathognomonic  of  putrid  infection,  is  denied  by  cer- 
tain experimenters.  The  same  may  be  said  of  the 
anatomical  lesions.  Thus  Gaspard,  Leuret,  Virchow, 
Panum,  Stick,  Henner,  Bergmann,  and  others  affirm 

*Rawitsch:  Zur  Lehre  von  der  putriden  Infection  und 
deren  Beziehungen  zura  sogenannten  Milzbrand.  Berlin,. 
1872. 


—    14  — 

that  hsemorrhagic  inflammation  of  the  intestinal 
mucosa  is  a  characteristic  lesion;  while  others  (Du- 
fiuis,  Sedillot,  Billroth,  J?aison,and  Schmidt)  have  found 
in  mortal  cases  the  intestinal  mucous  membrane 
intact.  Several  writers  think  that  sanguineous  ecchy- 
moses  in  the  different  organs  are  characteristic  of 
putrid  infection,  while  others  have  not  seen  them  in 
their  autopsies.  Bergmann  believes  that  the  modifi- 
cations of  the  spleen  are  constant  and  pathognomonic, 
while  Davaine  affirms  that  it  is  just  the  absence  of 
hypertrophy  of  the  spleen  in  putrid  intoxication  that 
enables  us  to  differentiate  it  from  charbon. 

To  explain  the  causes  of  these  variations  in  the 
results  of  experimentation,  writers  have  referred  us 
to  the  different  sources  of  the  putrid  substances, 
alleging  that  putrefied  meat  acts  differently  from 
putrefied  blood,  etc. — Stress  has  also  been  laid  on  the 
differences  of  the  action  of  the  putrid  poison,  accord- 
ing to  the  different  places  of  its  introduction  into  the 
animal  economy.  Thus,  the  effects  of  subcutaneous  in- 
jections were  distinguished  from  those  of  intravenous 
injection;  in  the  first  case,  a  more  or  less  intense 
local  inflammation  was  aroused,  complicated  with 
suppuration  and  gangrene;  in  the  second  case,  there 
appeared  the  septic  infection,  as  Gaspard  described 
it. — We  have,  moreover,  been  referred  to  the  different 
receptivity  of  the  animals  under  experimentation, 
varying  according  to  species,  race,  and  age. — But  it 
has  been  especially  remarked  that  the  toxicity  of 


putrid  products  undergoes  very  considerable  varia- 
tions in  the  course  of  putrefaction.  The  putrefied 
matters,  instead  of  becoming  more  and  more  toxic  as 
decomposition  went  on,  were  found  to  be  even  less 
toxic,  the  first  stages  only  giving  rise  to  the  most 
violent  poisons.  Billroth  shows  that  relatively  fresh 
pus,  even  when  it  is  not  putrid  but  "  good  and  laud- 
able," may  produce  by  intravenous  injection  very 
violent  and  febrile  reactions,  even  ending  in  death. 
H.  Fischer  demonstrated  that  pus  which  was  very 
toxic  at  the  onset,  loses,  in  putrefying,  its  specific  and 
septic  toxicity.  Bergmann  has  found  that  putrefied 
blood  is  more  toxic  in  the  first  five  to  six  days.  Heller 
has  remarked  the  same  fact. 

Samuel  has  studied  this  question  in  detail.  He 
finds  that  there  are  three  periods  in  the  toxicity  of 
putrid  substances:  phlogogenous,  septogenous,  and 
pyogenous.  In  the  first,  the  inoculation  of  putrid 
substances  is  followed  by  transient  inflammatory 
symptoms  which  disappear  without  leaving  any  local 
or  general  troubles.  In  the  septogenous  stage,  specific 
toxic  products  appear  which  provoke  the  true  "septic 
infection."  It  is  here  that  we  note  the  most  terrible 
local  and  general  phenomena  of  septicaemia:  the 
former  going  on  to  septic  gangrene,  the  latter  to  ful- 
minant septicaemia  which  kills  with  the  rapidity  of 
hydrocyanic  acid.  Later  on,  these  septic  properties 
of  the  progressively  decomposing  putrid  substances 
become  attenuated  little  by  little,  to  leave  remaining 


—  i6  — 

in  the  pyogenic  stage  only  the  power  of  prodncing 
localized  and  benign  suppurations. 

But  all  these  distinctions,  despite  their  consider- 
able interest,  did  not  quite  suffice  to  explain  the  in- 
constant and  conflicting  results  of  experimenters. 

There  remained  always  an  irreconcilable  differ- 
ence between  the  chemical  poison  of  Panum,  the  activ- 
ity of  which  depended  on  the  dose  injected,  and  the 
virus  of  Davaine,  which  acted  in  infinitesimal  doses  and 
which  was  found  in  increased  quantity  in  the  cadavers. 
It  was  evident  that  the  septicaemias  of  which  the  dif- 
ferent writers  spoke  were  not  at  all  identical,  and  that 
their  differences  depended  on  a  factor  having  quite 
other  powers  than  those  'alleged  by  the  chemists. 
But  above  all,  the  chemical  doctrine  was  unable  to 
explain  the  remarkable  fact  discovered  by  Coze  and 
Feltz,  and  confirmed  by  Davaine,  Hiller,  and  others: 
the  septic  poison,  instead  of  being  enfeebled  by 
dilution,  like  all  the  chemical  poisons,  in  the  cadavers 
of  the  animals  it  had  killed,  was  found  to  be  exalted. 
It  was  evidently  a  special  poison  fitted  to  reproduce 
itself  just  like  living  beings.  From  this  fact  there 
was  only  one  step  to  the  conclusion  that  this  poison 
was  constituted  by  bacteria.  One  might,  it  is  true, 
still  find  refuge  in  the  hypothesis  of  Liebig  and 
Robin,  that  the  soluble  ferments  are  albuminoid  mat- 
ters in  process  of  decomposition,  which  may  transmit 
their  alterability  to  other  bodies  and  reproduce  them- 
selves in  this  way  indefinitely.  But  this  hypothesis. 


of  Liebig  was  completely  ruined  in  its  chemical  ap- 
plications by  Pasteur,  who  demonstrated  peremptorily 
that  fermentation  (or  decomposition)  does  not  exist 
apart  from  the  life  of  organized  ferments.  Moreover, 
the  contagionists  were  soon  able  to  go  farther  in 
showing  the  fallacy  of  the  chemical  doctrine,  and  to 
prove  that  certain  microbes  separated  from  their 
morbid  products  were  able  to  produce  determinate 
diseases.  Thus,  it  was  proved  that  charbon,  hereto- 
fore ranked  among  the  putrid  fevers,  was  produced 
by  the  bacterium  of  Davaine;  that  septicaemia  was 
caused  in  man  by  the  septic  vibrio  of  Pasteur;  that 
the  different  septicaemias  and  pyaemias  in  animals  had 
each  its  special  pathogenic  agent,  living  staphylococci 
or  streptococci.  At  the  same  time  was  established 
the  fundamental  notion,  proclaimed  by  Ferdinand 
Cohn,  of  the  specificity  of  the  bacteria,  which  con- 
stitute each  a  being  apart,  characterized  by  its  mor- 
phology and  by  its  functions,  chromogenic,  zymotic, 
or  pathogenic.  Weigert  and  Ehrlich  brought  to  the 
aid  of  this  study  the  processes  of  staining  by  anilin 
dyes,  which  enable  microscopists  to  recognize  the  bac- 
teria. Koch  introduced  the  Abbe"  method  of  illumina- 
tion and  the  method  of  culture  on  solid  media,  which 
have  helped  bacteriologists  to  make  immense  strides 
in  their  knowledge  of  the  microbes.  Thus  was  con- 
stituted Bacteriology,  which  has  been  making  such 
rapid  advances.  It  is  now  known  with  certainty  that 
the  infectious  diseases  are  due  to  the  action  of  mi- 
crobes. 


—  i8  — 

The  vitalist  doctrine  was  not  in  absolute  contra- 
diction with  the  results  obtained  by  chemists,  and 
Panum  has  the  merit  of  having  understood,  just  as 
Selmi  and  Brieger  did  later,  the  possibility  of  recon- 
ciling the  two  theories. 

Panum  said  "that  it  is  incontestible  that  my 
putrid  poison  is  purely  chemical,  but  it  might  be  pro- 
duced by  a  microbe,  even  a  determined  microbe.  In 
diseases,  both  the  pathogenic  microbe  and  the  produc- 
tion of  the  poison  by  the  microbe  may  play  their 
respective  parts." 

But  the  conciliatory  voice  of  Panum  was  not 
heeded,  and  the  chemists  made  vigorous  opposition  to 
the  victorious  march  of  bacteriology.  They  continued 
to  bring  forward  proofs  in  favor  of  this  view — that  the 
microbes  were  but  simple  accompaniments  of  the 
poisons  of  Panum,  Bergmann,  and  Schmiedeberg. 
They  filtered  the  toxic  products  through  filtering 
paper  or  through  clay,  and  studied  the  effects  of  these 
filtered  liquids.  They  employed  heat,  dialysis,  the 
different  chemical  reagents,  to  eliminate  the  living 
microbes  of  their  experiments.  They  cultivated  the 
microbes  in  media  deprived  of  albuminoid  substances, 
to  show  that  in  these  cases  they  were  inoffensive 
(Hiller).  But  to  bacteriologists  all  these  experiments 
of  the  chemists  were  null  and  void,  for  they  could  not 
have  been  well  conducted  without  previous  knowledge 
of  the  properties  of  the  bacteria.  Nay,  more;  all  the 
experiments  on  systemic  intoxication  where  the  in- 


—  i9  — 

fluence  of  a  chemical  poison  made  itself  felt,  could 
but  embarrass  the  bacteriologists,  adding  a  supple- 
mentary factor  to  their  study  of  the  functions  of  bac- 
teria. 

Under  these  conditions  the  bacteriologists,  quite 
naturally,  made  a  profound  distinction  between  in- 
toxication and  infection.  Intoxication  was  character- 
ized by  the  sudden  appearance  of  the  accidents,  by 
the  proportionality  of  the  effects  to  the  doses  (doses 
always  considerable),  by  the  absence  of  specificity — for 
it  could  be  produced  by  the  most  common  bacteria  if 
injected  in  sufficient  quantity.  It  had  to  be  carefully 
avoided  in  bacteriological  studies.  These  concerned 
themselves  only  with  infection,  which  was  produced  by 
life  and  the  multiplication  of  specific  microbes  in  the 
animal  body.  It  was  independent  of  the  dose  of 
microbes  introduced  into  the  animal  body,  and  it  ap- 
peared after  an  incubation  necessary  for  their  multi- 
plication. The  infectious  disease  was  a  specific 
fermentation,  linked  to  the  life  and  multiplication  of 
the  pathogenic  microbes,  just  as  alcoholic  fermenta- 
tion depends  on  the  life  of  the  yeast. 

In  ignoring  intoxication  the  bacteriologists  had 
to  ignore  all  the  experimental  researches  on  the 
chemical  poisons;  and  the  dualism  of  intoxication  and 
infection  long  bore  as  a  heavy  weight  on  the  develop- 
ment of  microbiology.  But  during  this  time  the 
microbian  poisons  reappeared  in  a  quite  different 
science,  and  their  study  was  being  pursued  outside 
of  bacteriology. 


CHAPTER  III. 

THE  DISCOVERY  OF  PTOMAINES. 


SUMMARY. — Selmi  Draws  Attention  to  the  Ptomaine? 
Obtained  from  Cadavers — Leucoma'ines  and  Pto- 
maines of  Gautier — The  Researches  of  Brieger  on 
Ptomaines. 

In  a  celebrated  trial,  where  the  domestic  of  Gen. 
Gibbone  was  accused  of  the  death  of  the  latter,  the 
chemists  found  in  the  cadaver  the  alkaloid  delphinine. 
But  Francois  Selmi,  professor  of  chemistry  at  Bo- 
logne,  who  was  called  as  an  expert  witness  for  the 
defense,  demonstrated  by  a  detailed  chemical  analysis 
that  the  base  isolated  by  the  government  experts  was 
quite  distinct  from  the  delphinine  and  other  alkaloids 
of  vegetal  origin.  According  to  Selmi,  the  base  in 
question  was  of  animal  origin,  and  came  from  the 
putrefaction  of  the  cadaver.  This  dictum  of  Selmi 
was  completely  at  variance  with  the  general  convic- 
tion, according  to  which  the  alkaloids  could  only  be 
produced  by  plants.  Shortly  after,  similar  facts  were 
adduced,  and  drew  attention  more  and  more  to  the 
researches  of  Selmi.  In  the  trial  of  Sonzagno,  at  Cre- 
mona, the  first  experts  thought  they  had  found 
morphine  in  the  cadaver.  Selmi  peremptorily  demon- 


21    

strated  that  what  they  had  found  was  a  cadaveric 
alkaloid. 

There  was  still  later  a  celebrated  trial  in  Italy 
where  the  prosecution  endeavored  to  make  out  a  case 
of  poisoning  by  strychnine;  here  Selmi  affirmed  the 
cadaveric  source  of  the  base  found,  and  the  prisoner 
was  acquitted  on  this  testimony. 

Since  1870,  Selmi  has  been  making  experimental 
and  chemical  investigations  on  the  organic  bases 
found  in  cadavers.  To  this  order  of  researches  he 
was  led  by  noting  the  presence  in  cadavers  of  sub- 
stances which  possessed  the  general  and  even  the 
specific  reactions  of  alkaloids,  but  which  could  be 
distinguished  by  the  absence  of  all  toxic  action. 
Selmi  had  the  notion  that  alkaloids  similar  to  those  of 
animal  origin  could  be  produced  by  the  very  fact  of 
putrefaction.  To  verify  this  idea  he  instituted  a  long 
•series  of  researches  upon  exhumed  cadavers,  of  which 
the  disease,  the  death,  and  the  duration  of  burial  were 
known  to  him.  He  there  discovered,  in  fact,  a  great 
number  of  both  inoffensive  and  toxic  alkaloids,  with 
different  chemical  properties,  and  more  or  less  like 
the  alkaloids  of  plants. 

Pursuing  further  his  researches  on  these  products, 
Selmi  demonstrated  their  origin  in  the  putrefaction  of 
albuminoid  matters,  for  he  found  that  ptomaines  also 
take  rise  in  the  albumin  of  putrefied  egg. 

These  discoveries  of  Selmi  were  too  revolutionary 
of  former  notions  to  be  accepted  without  opposition. 


22    

We  have  already  mentioned  how,  in  certain  criminal 
prosecutions,  they  forced  themselves  upon  the  atten- 
tion of  the  whole  scientific  world. 

These  expert  findings  of  Selmi  in  Italy  had  also 
their  counterpart  in  Germany,  in  the  celebrated  trial 
of  Brandes-Klebs  in  1874.  Chemists  had  extracted 
from  the  cadaver  of  Klebs  a  very  toxic  alkaloid  which 
they  had  identified  with  coniine.  Otto  showed  that 
it  was  neither  coniine  nor  nicotine,  though  it  resembled 
these  alkaloids.  He  believed  it  to  be  a  ptomaine,, 
and  this  was  the  opinion  of  the  experts. 

It  is  doubtless  true  that  Selmi  was  not  the  first  to 
find  organic  bases  of  animal  origin;  that  Marquardt, 
of  Stettin,  in  1865  extracted  from  a  human  cadaver  a 
toxic  alkaloid  resembling  coniine,  which  he  called 
septicine;  that  Dupr£  and  Bence  Jones,  in  1868, 
found  in  animal  bodies  a  substance  bearing  a  marked 
resemblance  to  an  alkaloid;  that  Sonnenschein,  in 
1869,  discovered  in  a  cadaver  an  alkaloid  which  he 
thought  was  coniine,  but  which  proved  not  to  be  toxic; 
the  merit  none  the  less  belongs  to  Selmi  of  having  been 
the  first  to  subject  these  animal  bases  to  a  profound 
series  of  researches,  and  of  thereby  explaining  their 
true  meaning  and  importance.  It  is  noteworthy  that 
in  Selmi's  estimation  the  interest  in  the  study  of 
ptomaines  was  by  no  means  limited  to  legal  medicine 
by  reason  of  their  simulation  of  vegetal  alkaloids. 
He  had  well  comprehended  the  signification  which 
they  might  have  for  biological  chemistry  and  physio- 


—  23  — 

logical  pathology.  In  biological  chemistry  the  pto- 
maines were  to  Selmi  a  new  demonstration  of  the 
absence  of  any  clear-cut  limits  between  the  vegetable 
and  animal  kingdoms.  At  the  expense  of  animal 
matters  might  be  produced  alkaloids  quite  like  those 
which  down  to  Selmi's  time  were  believed  to  be  exclu- 
sively of  vegetal  origin.  To  general  pathology  the 
importance  of  the  discoveries  of  Selmi  was  still 
greater.  The  same  substances  which  the  microbes 
may  produce  at  the  expense  of  the  dead  animal  body, 
they  may  elaborate  in  the  living  body;  in  other 
words,  the  infectious  disease  produced  by  the  microbes 
may  give  rise  to  the  formation  of  ptomaines  in  the  dis- 
eased organism.  In  fact,  Selmi  found  ptomaines  in 
the  urine  in  cases  of  typhoid  fever,  pneumonia,  and 
tetanus.  The  premature  death  of  Selmi  arrested  these 
important  researches. 

The  movement  imparted  to  science  by  Selmi  was 
of  very  great  importance.  A  large  number  of  memoirs 
have  been  recently  published  on  the  ptomaines.  To 
mention  only  a  few,  Nencki  in  1876  gave  the  first 
analysis  of  a  ptomaine,  obtained  by  the  putrefaction 
of  gelatin  with  the  pancreas.  This  was  collidine,  a 
base  already  extracted  from  the  products  of  distilla- 
tion of  animal  oil. 

Morrigia  and  Battistini  in  1875  showed  that  very 
toxic  ptomaines  can  be  got  from  cadavers,  whose  action 
is  quite  similar  to  that  of  curare.  Rorsch  and  Fass- 
bender  found  in  the  cadaver  a  ptomaine  resembling 


—   24  — 

digitalin  in  its  action.  Brouardel  and  Bontmy  dis- 
covered a  base  very  similar  to  veratrine.  Gautier 
and  Etard,  in  1881,  defined  chemically  two  other 
bases,  parvoline  and  hydrocollidine,  found  in  the 
products  of  decomposition  of  fish,  and  of  the  meat  of 
the  horse  and  the  ox.  The  fourth  ptomaine  defined 
was  corindine,  found  in  1883  by  Quareschi  and  Mosso. 

We  must  devote  more  time  to  the  extremely  im- 
portant researches  of  Armand  Gautier. 

Gautier  had  announced  in  1871  that  during  the 
putrefaction  of  fibrin  there  is  formed  a  certain 
quantity  of  alkaloids,  both  fixed  and  volatile.  In 
1881-1883  he  made,  with  Etard,  a  series  of  researches 
which  disclosed  the  chemical  nature  of  certain  pto- 
maines of  putrefaction  belonging  to  the  pyridic  and 
hydropyridic  series. 

According  to  Gautier,  all  the  decompositions  of 
albuminoid  bodies  taking  place  away  from  the  air 
give  rise  to  alkaloids.  Putrefaction  is  always  accom- 
panied by  the  formation  of  ptomaines;  but  the  normal 
life  of  animals,  which  in  great  part  goes  on  in  the  ab- 
sence of  free  oxygen,  leads  also  to  the  formation  of 
organic  bases,  which  Gautier  has  grouped  under  the 
name  of  leucomaines.  Such  are  xanthine,  creatine, 
etc.  Whenever  perturbations  of  the  normal  con- 
ditions of  life  augment  the  want  of  oxygen  in  the 
body,  as  in  anaemia  and  the  infectious  diseases,  then 
the  basic  products  or  ptomaines  arise  in  increased 
quantity.  The  non-elimination  of  the  normal  leuco- 


—  25  — 

maines,  and  the  production  ol  ptomaines,  are  so  many 
Causes  of  intoxication  (or  poisonjnp^  of  the 


Certain  leucomaines  are  extremely  toxic;  for  instance, 
the  secretions  of  the  different  venomous  animals. 
(Already,  in  1866,  Zalewsky  had  isolated  from  the 
salamander  the  alkaloid  samandrinej) 

We  have  seen  with  what  amplitude  of  view  Gau- 
tier  established  his  conception  as  to  the  origin,  virtu- 
ally identical,  of  the  alkaloids  in  vegetables,  animals, 
and  microbes.  These  bases  of  the  same  nature  are 
derived  everywhere  from  the  same  source,  the  fllhij- 
mins;  everywhere  in  the  same  fundamental  condi- 
tions^—d  ^2^£^M!?£.  JjlL^tfaCL  .absencfer  ^£— g^no^ 
quantity  of  oxygen.  Guided  by  these  general  ideas, 
Gauffer  and  his  pupils  found  a  great  number  of  new 
leucomaines.  But  the  exposition  of  their  history  does 
not  enter  into  the  plan  of  this  book. 

The  study  of  the  ptomaines  properly  so  called, 
which  interest  us  particularly  here,  owes  its  great 
progress  during  the  past  few  years  to  the  researches 
of  Brieger.  Brieger  has  made  a  systematic  study  of  a 
great  number  of  questions  called  up  by  the  history  of 
the  ptomaines.  Thanks  to  the  perfected  chemical 
methods,  he  has  succeeded  in  isolating  numerous 
ptomaines  perfectly  denned  as  chemical  units.  We 
shall  indicate  the  principal  results  which  he  obtained. 
Seeing  the  facility  with  which  the  ptomaines  form  in 
the  midst  of  albuminoid  substances  in  putrefaction, 
Brieger  asked  if  the  alkaloids  might  not  result  from 


—     26    — 

the  simple  solution  of  the  albumins  by  the  soluble 
ferments.  In  fact,  in  digesting  meat  by  pepsin,  he 
noted  the  appearance  among  the  products  of  this 
digestion  of  a  convulsivant  organic  base,  which  he 
called  peptotoxine.  We  shall  return  later  to  this 
remarkable  result,  which  has  an  enormous  importance 
in  connection  with  this  question  of  ptomaines. 

In  putrid  cadavers,  and  in  the  flesh  of  mammals 
and  of  putrid  fish,  Brieger  found  a  certain  number  of 
bases  regarded  by  him  as  constant  representatives  of 
the  ptomaines  of  putrefaction.  They  all  belong  to 
the  category  of  ammoniacal  compounds,  amines.  Be- 
sides the  methyl  and  ethyl  amines,  Brieger  has  isolated 
in  a  state  of  perfect  purity  cadaverine,  putrescine, 
neurine,  muscarine,  ethylendiamine,  saprine,  myda- 
leine,  and  a  great  number  of  others. 

They  are  chiefly  diamines.  Lastly,  Brieger  took 
up  again  the  idea  of  Panum  and  of  Selmi,  and  made 
a  study  of  the  ptomaines  produced  by  the  microbes 
of  infectious  diseases.  He  has  studied  also  the  bases 
formed  in  the  cultures  by  the  typhic  and  tetanic 
bacilli,  by  the  vibrio  of  cholera.  But  by  these  latter 
studies  Brieger  enters  into  the  new  period  of  the 
evolution  of  ideas  concerning  the  rdle  of  bacterial 
poisons,  a  period  which  shall  be  set  forth  in  the  fol- 
lowing chapter. 

A.  On  the  Constitution  of  the  Artificial  and 
Natural  Alkaloids. 

The  first  vegetable  alkaloid,  morphine,  was  dis- 


_  27  — 

covered  by  Sertiirner  in  1806.  Since  then  the  num- 
ber of  organic  bases  extracted  from  plants  or  pre- 
pared artificially  has  been  considerable,  especially 
during  the  past  few  years,  and  chemists  have  at  last 
succeeded  in  determining  their  constitution. 

The  principal  alkaloids  are  derivatives  of  pyri- 
dine,  which  much  resembles  benzol. 

Pyridine  is  a  benzol  where  an  atom  of  nitrogen 
(N,  which  is  trivalent)  replaces  the  group  CH  (which 
has  also  three  free  atomicities). 

In  the  formula  of  pyridine,  the  atoms  of  hydrogen 
may  be  replaced  by  the  monovalent  groups — 

C 

H-  |  -H,  or  (CH.) 
H 

Thus  we  have  picoline.  If  this  substitution  be  made 
twice,  we  shall  have  putidine;  with  three  groups  of 
CH3  instead  of  3  atoms  of  H,  we  obtain  collidine, 
and  after  a  new  substitution,  corindine. 

All  these  products  are  energetic  bases  obtained 
artificially  by  the  dry  distillation  of  bones  (oil  of 
Dippel).  Collidine  and  corindine  are  already  known 
to  us,  for  they  have  also  been  found  in  the  products 
of  putrefaction. 

The  natural  alkaloids  of  plants  are  a  little  more 
complex,  but  they  are  also  the  substitution  products 
of  pyridine  or  of  piperidine  (which  is  hydrogenated 
pyrodine). 

Thus,  coniine  is  a  propylpiperidine,  or  a  piperidine 


—    28    — 

where  an  atom  of  H  is  replaced  by  the  monoatomic 
group  C3H7  which  is  called  propyl.  Other  alkaloids 
of  plants  are  derivatives  of  quinoline,  which  corre- 
sponds to  naphthaline  where  an  atom  ojE.  N  replaces 
the  group  CH.* 

B. —  The  Amines. 

Ammonia  has  the  formula: 

— H 

NH8,  or  N— H. 
— H 

The  atoms  of  H  in  ammonia  may  be  replaced  by  the 
alcohol  radicals.  This  substitution  gives  rise  to  the 
ammoniacal  compounds,  or  the  amines. 

Thus,  methyl-alcohol  has  the  formula  CH3HO. 
In  this  formula  CH3  is  the  alcohol  radical  called 
methyl,  combined  with  the  group  HO,  which  is  called 
hydroxyle.  The  methyl  may  replace  one,  two,  or 
three  atoms  of  hydrogen  in  the  ammonia.  We  shall 
have  monomethylamine,  dimethylamine,  trimethyl- 

amine: 

— H                     — H  — CH8 

N— H  N— CH8  N— CH3 

— CH8                  — CH8  — CH8 

The  bivalent  alcohol-radicals  may  unite  with  two 
molecules  of  ammonia.  The  resulting  bodies  are 
called  diamines.  Thus,  ethylen — C2H4 — which  has 
the  two  atomicities  free,  forms  ethylendiamine: 

-C8H4 
N— HH— N 
— HH— 

*  Von  Pictet:  The  Chemical  Constitution  of  the  Vegetal 
Alkaloids.  Paris,  1888. 


—  29  — 

Most  of  Brieger's  ptomaines  are  diamines.  Thus, 
putrescine  is  a  tetramethylendiamine;  the  bivalent 
alcohol-radical  CH2  is  called  methylen: 

— H  H 

N— CH8— CH8— CH2— CH8— N 
-H  H 

Cadaverine  is  a  pentamethylendiamine: 

— H  H 

N— CH2— CH8— CH2— CH2— CH2— CH8— N 
— H  H 

REFERENCES. 

Selmi:  "Ptomaine  od.  Alcaloide  Cadaverice."  Bo- 
logna, 1881. 

Husemann:  "Die  Ptomaine  und  ihre  Bedentung  fttr 
die  Gerichtliche  Chemie  und  Toxicologie "  (Archiv  der 
Pharmacie,  Bd.  16,  17,  19,  20,  and  21). 

(Effinger:  "  Die  Ptomaine  oder  Cadaver  Alcaloide." 
Wiesbaden,  1885. 

Gautier:  "  On  the  Alkaloids  Derived  from  the  Destruc- 
tion of  Animal  Tissues,  Ptomaines  and  Leucomalnes  "  (Bull, 
de  1'Acad.  de  M6d.,  Jan.  12  and  16,  1886). 

Gautier:  "  Chimie  Biologique."     Paris,  1892. 

Hogouneq:  "  The  Alkaloids  of  Animal  Origin."  Paris, 
1886. 

Roussy:  "  Ptomaines  and  Leucomalnes  "  (Rev.  des  Sc. 
Med.,  t.  xxxi,  1888). 

Brieger:  "  Untersuchungen  uber  Ptomaine."  Three 
volumes.  Berlin,  1885-1886. 

Brieger:  "  Beitrag  zur  Kentniss  der  Zusammensetzung 
des  Mytilotoxins  nebst  einer  Uebersicht  der  Bisher  in  ihrer 
Haupteigenschaften  bekannten  Ptomaine  und  Toxine '" 
(Virchow's  Archiv  f.  Pathol.  Anat.,  1889,  t.  115,  p.  483). 


CHAPTER  IV. 

INFECTION    IS  AN    INTOXICATION    BY   THE    Ml- 
CROBIAN    POISON. 


SUMMARY. — Bacteriology  Aims  to  Explain  Infection  by 
the  Invasion  of  the  Animal  Economy  by  Microbes — 
Three  Diseases,  Diphtheria,  Cholera,  Tetanus, 
where  this  Explanation  is  Not  Possible — Discovery 
of  Chemical  Vaccination — Discovery  of  the  Different 
Chemical  Poisons  of  Bacteria  which  May  Produce 
All  the  Symptoms  of  the  Disease  without  the  Inter- 
vention of  Microbes — Actual  Point  of  View:  The 
Infectious  Disease  is  an  Intoxication  by  the  Poison 
of  the  Pathogenic  Microbe. 

We  quit  bacteriology  at  the  moment  when,  obtain- 
ing a  decisive  victory  over  the  chemical  etiology  of 
the  infectious  diseases,  it  denied  the  importance  of 
the  poisons  produced  by  the  microbes.  The  faculty 
of  the  bacteria  of  fabricating  poisons  presented  no 
interest  to  the  bacteriologists,  for  it  belonged  to  the 
common  bacteria,  those  of  putrefaction,  which  live  on 
materials  deprived  of  life,  while  the  pathogenic  bac- 
teria were  characterized  by  their  power  to  develop  in 
the  living  organism  and  to  invade  it. 

We  must  remember  that  bacteriology  originated 


in  the  study  of  fermentation,  with  which  naturally 
the  infectious  disease  came  to  be  assimilated.  As 
alcoholic  fermentation  had  found  its  provisional  ex- 
planation in  the  life  without  air  of  the  yeast  plant, 
so  the  disease  was  explained  by  the  life  of  the  microbe 
in  the  animal  body. 

Thus,  for  example,  in  the  deadly  anthrax,  whose 
study  has  been  intimately  associated  with  the  begin- 
nings of  bacteriology,  and  which  long  served  as  the 
type  of  the  bacterial  diseases,  the  animals  died  with  a 
great  quantity  of  bacteria  in  the  blood  of  all  the 
organs.  It  was  suppesed  that  the  death  of  the  ani- 
mals was  caused  by  this  enormous  mass  of  foreign 
bodies  invading  the  animal  humors,  and  the  mechan- 
ism of  the  disease  was  explained  by  the  mechanical 
effects  of  the  presence  of  these  foreign  bodies  in  the 
blood,  and  the  capillary  emboli  therefrom  resulting. 
Writers  also  insisted  on  the  evils  resulting  from  the 
subtraction  of  oxygen  from  the  red  globules  by  the 
anthracoid  bacteria,  the  lack  of  oxygen  in  the  tissues, 
and  the  asphyxia  therefrom  resulting.  They  also  had 
much  to  say  about  the  asphyxia  of  the  animal  by  the 
carbonic  acid  set  free  by  the  microbes. 

In  a  more  general  way  they  directed  our  atten- 
tion to  the  energetic  nutrition  of  the  bacteria,  which 
get  possession  of  the  pabulum  of  the  animal  cells  and 
kill  them  when  starved. 

Oftener  still  they  contented  themselves  with 
vague  expressions  about  the  struggle  for  existence 


—  32  — 

between  the  animal  cell  and   the   microbe,  without 
attempting  further  to  analyze  this  vast  conception.* 

This  was,  we  repeat,  because  the  rising  bacteri- 
ology was  dominated  by  the  analogy  between  the  in- 
fectious disease  and  fermentation  by  figured  bodies. 
Just  as  the  decomposition  of  matters  in  fermentation 
was  produced  not  by  a  chemical  substance,  or  enzyme, 
but  by  the  life  without  air  of  the  ferments,  so  the 
infectious  disease  was  the  corollary  of  the  life  of 
the  pathogenic  bacteria  in  the  animal  body.  And  we 
have  just  seen  that  absorption  of  oxygen  by  the  bac- 
teria was  especially  invoked  in  explanation  of  the 
mechanism  of  the  disease. 

At  the  same  time,  the  idea  of  intoxication  by 
chemical  poisons,  which  at  an  early  period  dominated 
infection,  was  not  completely  given  up.  Davaine  be- 
lieved that  the  anthracoid  bacteria  secrete  a  product 
which  agglutinates  the  red  globules  of  the  blood. 
Pasteur,  by  the  filtration  of  anthracoid  cultures,  proved 
the  existence  of  this  agglutinating  diastase.  Toussaint 
invoked  the  existence  of  a  phlogistic  poison  produced 
by  the  anthracoid  bacteria.  He  believed  that  he 
could  even  isolate  it  and  confer  with  it  immunity 
against  charbon.  Chauveau  also  adduced  arguments 
in  favor  of  this  chemical  vaccination  by  the  bacterial 
poisons. 

But  chemical  vaccination  found  a  formidable  op- 


*Duclaux:  "  Le  Microbe  et  la  Maladie."     Paris,  1866. 


—  33  — 

ponent  in  Pasteur,  who  saw  in  it  the  overthrow  of  his 
vitalist  doctrine  of  fermentation  and  disease.  As  in 
the  fermentations  the  diastases  or  soluble  ferments 
play  only  a  secondary  role  in  preparing  the  substances 
for  the  principal  transformations  effected  by  the  living 
ferments,  so  the  diastases  of  the  pathogenic  bacteria 
can  only  serve  to  explain  the  relatively  unimportant 
symptoms  of  the  disease  which  itself,  as  well  as  the 
chemical  vaccine,  was  due  solely  to  the  life  of  the 
bacteria. 

But  in  proportion  as  bacteriology  made  its  new 
conquests,  the  insufficiency  of  this  pathogeny  of  the 
infectious  diseases  became  the  more  manifest. 

In  1884,  Loffler  found  the  bacillus  of  diphtheria. 
He  showed  that  this  bacillus  is  always  strictly  localized 
in  man  to  the  mucous  membranes  which  are  the  seat  of 
the  lesion.  So  in  animals  we  only  find  it  at  the  site 
of  inoculation,  and  not  in  the  internal  organs,  where 
it  cannot  live. 

This  microbe,  which  does  not  invade  the  organ- 
ism, is  nevertheless  very  pathogenic.  Loffler  showed 
that  this  pathogenic  action  is  due  to  the  production 
by  the  bacillus  of  a  violent  chemical  poison,  which  is 
absorbed  at  the  place  where  the  bacillus  makes  its 
habitat  and  vegetates. 

The  problem  of  microbian  poisons  still  more  im- 
peratively urged  itself  on  the  attention  of  the  profes- 
sion when  Koch  discovered  his  cholera- vibrio. 

Koch  demonstrated  that  the  comma  bacillus  does 
3  ooo 


—  34  — 

not  penetrate  the  internal  organs  of  the  victims  of 
cholera,  but  remains  always  limited  to  the  intestinal 
canal.  It,  however,  produces  the  grave  symptoms  of 
cholera  poisoning.  Koch  concluded  that  the  comma 
bacillus  secretes  a  violent  poison  which  explains  the 
clinical  phenomena  of  the  disease.  His  prevision 
turns  out  to  be  supported  by  experiment.  Cholera 
cultures  sterilized  in  totality,  or  filtered  of  bacteria, 
and  even  the  ptomaines  extracted  from  these  cultures, 
have  been  found  endowed  with  a  toxicity  more  or 
less  appreciable. 

A  third  bacterial  disease  was  soon  found  to  be 
absolutely  inexplicable  without  the  intervention  of  a 
soluble  poison.  This  is  tetanus.  The  bacillus  of 
tetanus,  discovered  by  Nicolaier,  is  strictly  limited  to 
the  place  of  its  inoculation.  It  nevertheless  produces 
a  terrible  constitutional  intoxication.  Brieger  sought 
for  the  tetanic  poison,  and  found  in  the  cultures  of 
the  bacillus  and  in  the  tetanized  muscles  several 
specific  ptomaines  which  provoked  convulsions  in 
animals. 

The  great  importance  of  the  microbial  poisons 
was  emphatically  demonstrated,  thanks  to  the  triumph 
of  the  doctrine  of  chemical  vaccines. 

The  possibility  of  vaccinating  animals  against 
infection  by  the  soluble  products  of  bacteria,  affirmed 
in  the  case  of  charbon  by  Toussaint  and  by  Chauveau, 
and  denied  by  Pasteur,  was  established  several  years 
later  by  the  researches  of  numerous  experimenters — 


—  35  — 

e.g.,  Wooldridge  for  charbon,  Salmon  and  Smith  for  hog 
cholera,  Beumer  and  Peiper  for  typhoid  fever,  Char- 
rin  for  the  pyocyanic  disease.  Latterly  researches  on 
chemical  vaccinations  have  multiplied,  and  we  may 
without  exaggeration  affirm  that  the  acquisition  of 
immunity  in  infectious  diseases  is  a  fact,  thanks  to 
the  chemical  products  of  the  microbe.  Lastly,  we 
have  succeeded  in  separating  the  microbes  from  their 
chemical  poisons  by  filtration. 

The  decisive  step  in  this  direction  was  taken  by 
Roux  and  Yersin.  They  found  that  after  filtering 
through  the  Chamberland  filter  the  cultures  of  the 
diphtheria  bacillus,  they  obtained,  with  the  filtered 
liquid  entirely  deprived  of  bacilli,  toxic  effects  on  the 
animals  under  experimentation.  These  toxic  effects 
are  obtained  with  minimum  doses,  and  resemble  the 
infection  after  incubation  or  after  inoculation  with 
the  living  bacilli.  We  may,  for  example,  produce 
by  the  injection  of  filtered  cultures  the  same  diph- 
theritic paralyses  as  with  the  bacillus  itself.  This 
identity  of  effects  constitutes  a  real  demonstration  of 
Loffler's  notion  that  the  diphtheritic  bacilli  act  on  the 
animal  economy  by  the  poison  which  they  produce. 

The  researches  of  Roux  and  Yersin  were  soon 
confirmed  by  Loffler  himself  and  by  many  other 
experimenters. 

It  was  also  found  that  diphtheria  is  not  the  only 
disease  producible  by  the  chemical  products  of  the 
specific  micro-organism.  Filtered  cultures  of  the 


-  36- 

bacilli  of  tetanus  were  found  by  Knud-Faber,  Tiz- 
zoni,  Vincent,  and  others,  to  be  extremely  toxic. 
These  filtration  products,  injected  in  animals,  re- 
produced in  them  all  the  symptoms  so  characteristic 
of  tetanus.  Lastly,  in  the  case  of  cholera  also,  I  have 
succeeded  in  reproducing  all  the  typical  symptoms  of 
the  disease  with  the  cultures  of  Koch's  vibrio  steril- 
ized by  discontinuous  heat.  From  many  other  microbes, 
as  we  shall  see  as  we  go  on,  poisons  more  or  less  viru- 
lent have  been  separated. 

At  the  onset  of  this  period  of  the  discovery  of 
bacterial  poisons,  it  was  believed  that  we  should  have 
to  distinguish  two  classes  of  infectious  diseases.  In 
one  class  would  be  ranged  the  infectant  microbes 
which  produce  the  disease  and  kill  solely  by  their 
abundant  development  in  the  body  of  the  animal; 
as  types  of  this  form,  we  were  referred  to  charbon, 
tuberculosis,  pneumonia,  septicaemia  in  the  hare. 
These  microbes  were  supposed  not  to  have  the  power 
to  fabricate  poisons.  In  the  second  class  we  had  the 
toxine-making  microbes,  as  those  of  cholera,  tetanus,, 
diphtheria,  which  cannot  pullulate  abundantly  in  the 
animal  economy,  but  are  endowed  with  extreme  toxi- 
genous  power. 

But  this  distinction  is  not  legitimate.  First,  the 
same  microbes  produce  in  certain  animal  species  gen- 
eral septicjmaladies,  and  local  lesions  in  other  species: 
the  anthracoid  bacterium  and  the  lanceolate  strepto- 
coccus of  pneumonia  are  examples.  A  still  more  im- 


—  37  — 

portant  argument  against  this  distinction  is  that  in 
the  case  of  the  so-called  infectant  microbes,  we  have 
succeeded  in  finding  the  poisons  which  they  prepare. 
Thus  the  anthracoid  poison  has  been  described  by 
Martin  and  Christmas;  the  pneumonic  poison  by  G. 
-and  F.  Klemperer;  the  tuberculous  poisons  by  Koch, 
Maffuci,  Prudden  and  Hoddenpyl,  Strauss,  and  my- 
self. 

By  all  the  researches  which  we  have  reported  in 
this  chapter,  it  has  been  established  to  a  certainty  that 
the  pathogenic  microbes  act  on  the  animal  organism 
by  means  of  the  poisons  which  they  produce.  From 
this  point  of  view  the  importance  of  the  microbian 
poisons  becomes  extremely  great. 

At  the  outset  of  our  historical  survey  we  saw  that 
systemic  intoxication,  to  Hiller  and  his  partisans, 
dominated  infection;  the  poisons  "  formed  in  the  midst 
of  the  tissues  in  decomposition"  were  everything, 
and  the  microbes  nothing,  in  the  disease. 

Later  the  importance  of  these  two  factors  was 
inverted:  the  intoxication  was  a  common  place  phe- 
nomenon, meriting  no  special  attention;  the  entire  dis- 
ease was  produced  by  the  progressive  invasion  of  the 
living  animal  by  the  microbe. 

To  day  intoxication  has  reconquered  its  rights. 
Infection  is  considered  as  an  intoxication,  but  it  is  a 
special  intoxication  by  the  specific  poison  of  a  patho- 
genic bacterium. 

Progress,  as  Hegel  has  defined  it,  traverses  three 


_  38  - 

successive  stages.  The  second  is  the  negation  of  the 
first.  The  last  approaches  the  first  in  its  form,  but 
explains  the  contradiction  of  the  first  two  phases. 


Second  Part:   General  Microbial  Toxi- 
cology. 

CHAPTER  V. 

THE   CHEMICAL    NATURE    OF   THE    BACTERIAL 
POISONS.' 


SUMMARY. — Different  Conceptions  of  the  Chemical  Na- 
ture of  the  Bacterial  Poisons.  First  Stage:  The 
Ptomaines — Differences  between  the  Ptomaines  of 
Selmi,  of  Gautier,  and  of  Brieger — The  Pepto- 
toxine  of  Brieger — Researches  of  Salkowsky,  of 
Bouveret  and  Devic — Researches  of  Bassi  on 
Typhotoxine — Work  of  Baumann — The  Ptomaines 
are  Perhaps  Artificial  Products — Opinion  of  S. 
Martin.  Second  Stage:  The  Diastases — Researches 
of  Roux  and  Yersin — Criticism  of  the  Notion  of 
Diastases.  Third  Stage:  The  Toxalbumins — 
Work  of  Brieger  and  Frdnkel. 

The  conceptions  of  the  bacteriologists  on  the 
chemical  nature  of  the  bacterial  poisons  have  already 
gone  through  several  successive  stages,  under  the  in- 
fluence principally  of  other  toxicological  researches 
of  the  same  epoch. 


—  40  — 

Panum,  whose  brilliant  achievements  deserve  so 
often  to  be  cited,  had  found  that  the  principal  poison 
of  putrid  matters  is  insoluble  in  alcohol,  and  re- 
sembles certain  albuminoid  substances  which,  like 
the  peptones,  are  not  modified  by  boiling.  But  since 
Panum's  researches,  the  attention  of  investigators  has 
been  directed  more  and  more  to  toxic  substances 
which  are  soluble  in  alcohol  and  are  analogous  to  the 
most  formidable  of  known  poisons,  the  vegetal  alka- 
loids. 

Of  the  putrid  poison  of  Gaspard  and  of  Panum, 
Selmi,  Gautier,  and  Brieger  studied  only  one  part, 
the  ptomaines.  Regarding  their  task  from  a  purely 
chemical  point  of  view,  these  three  chemists  bestowed 
but  little  study  on  the  physiological  action  of  the 
toxic  substances  they  were  testing.  Nor  did  they 
compare,  as  Panum  did,  the  toxicity  of  the  pri- 
mary poison  with  that  of  the  different  extracts  ob- 
tained from  it.  The  sole  end  of  Selmi,  Gautier,  and 
Brieger  was  to  extract  from  the  putrid  matters  in  a 
state  of  perfect  purity  certain  definite  chemical  bodies, 
the  ptomaines,  whose  pre-existence  in  these  matters 
did  not  seem  to  them  to  merit  a  special  demonstra- 
tion. All  three  have  succeeded  in  their  efforts. 

But  it  would  be  very  instructive  to  enter  a  little 
more  into  detail  in  the  examination  of  the  results 
which  they  have  obtained,  and  to  compare  the  differ- 
ent ptomaines  of  these  three  writers. 

Selmi  found  some  cadaveric  alkaloids  extremely 


like  those  of  vegetal  origin.  Thus,  for  instance,  he 
has  isolated  a  cadaveric  coniine,  which  by  all  its 
chemical  reactions  and  by  its  physiological  action  is 
not  distinguishable  from  the  vegetal  alkaloid. 

The  other  chemists,  contemporaries  of  Selmi, 
found  also,  as  we  have  already  seen,  ptomaines  re- 
markably like  the  veritable  alkaloids.  We  need  only 
mention  the  bases  which  resemble  conicine  (Sonnen- 
schein),  nicotine  (Wolkenhaar),  atropine  (Tuelzer), 
veratrine  (Brouardel  and  Bontmy). 

Later  Gautier,  Nencki,  Quareschi,  and  Mosso 
found  other  ptomaines  which  constantly  accompany 
putrefaction.  These  were  collidine,  parvoline,  corin- 
dine — bodies  less  complex  than  the  vegetal  alkaloids, 
but  still  belonging  to  the  pyridic  series. 

Later  still,  Brieger  studied  putrefaction  under  its 
various  aspects.  He  discovered  everywhere  new 
ptomaines,  but  did  not  succeed  in  finding  those  of 
Selmi,  nor  even  those  of  Gautier.  The  ptomaines  of 
Brieger  are  diamines,  and  do  not  belong  to  the  pyridic 
series. 

To  what  can  these  constant  differences  be  due, 
and  why  have  these  chemists  constantly  found  these 
different  products? 

Evidently,  we  must  hold  in  suspicion  their  differ- 
ent methods  of  extraction  of  ptomaines.  We  might 
even  be  warranted  in  believing  that  these  methods 
were  likely  to  create  artificial  products  which  did  not 
pre-exist  in  the  matters  analyzed.  We  cannot  here 


—  42  — 

enter  into  the  chemical  details  as  to  the  methods  em- 
ployed by  Selmi,  Gautier,  and  Brieger;  we  will  con- 
tent ourselves  with  the  analysis  of  only  one  special 
point  of  this  question.  We  will  study  the  history  of 
the  peptotoxine  of  Brieger.  This  history  has  a  capital 
importance  in  connection  with  the  entire  question  of 
ptomaines  and  their  origin  in  albuminoid  matters. 

We  have  already  seen  (see  Chap/ III)  that  Brieger 
found  a  toxic  ptomaine  among  the  products  of  peptic 
digestion.  In  subjecting  the  different  species  of  albu- 
min, or  even  of  peptone,  to  digestion  with  gastric  juice, 
Brieger  discovered  a  toxic  body  soluble'  in  ethyl  and 
amyl  alcohol,  giving  the  characteristic  reaction  of  the 
alkaloids.  Brieger  gave  the  name  of  peptotoxine  to 
this  ptomaine,  which  he  supposed  to  be  an  aromatic 
amine.  As  all  albuminoid  matters  in  undergoing  de- 
composition begin  by  being  converted  into  peptones, 
it  is  quite  natural  that  Brieger  should  find  his  pepto- 
toxine at  the  onset  of  all  the  putrefactions  which  he 
studied.  It  disappeared  after  the  first  week.  We  see 
what  great  interest  is  attached  to  peptotoxine.  This 
toxic  ptomaine  is  the  necessary  stage  through  which 
pass  all  the  albumins  in  being  liquefied  by  the  soluble 
ferments  of  animal  or  bacterial  origin.  Being  formed 
at  the  expense  of  the  albumins  by  the  relatively  simple 
mechanism  of  diastasic  fermentation,  peptotoxine 
might  well  serve  as  a  key  to  explain  the  appearance 
of  ptomaines  with  the  intervention  of  microbes. 

Salkowsky,  who  has  studied  with  much  care  this 


—  43  — 

question  of  the  peptotoxine  of  Brieger,  has  never  been 
able  to  find  it  in  the  products  of  gastric  digestion. 
He  supposed  that  Brieger's  alkaloid  was  produced  by 
the  intervention  of  putrefaction  during  and  with  the 
aid  of  the  manipulations.*  The  question  has  just  re- 
ceived an  unexpected  solution  by  the  researches  of 
Bouveret  and  Devic.  f  These  writers  have  demon- 
strated that  peptotoxine  is  an  artificial  product  which 
is  formed  at  the  expense  of  albuminoid  matters  by  the 
combined  action  of  hydrochloric  acid  and  of  alcohol. 
Already  Tanret  had  insisted  on  the  close  kinship 
between  the  peptones  and  the  alkaloids.  He  had 
even  shown  that  by  the  action  of  soda  on  the  peptones, 
we  may  form  alkaloids  extractable  by  ether.  Later, 
Dreschel  saw  that  albuminoid  substances  boiled 
with  acids  yield  among  the  products  of  their  de- 
composition the  organic  bases.  Bouveret  and  Devic 
have  now  shown  that  even  less  energetic  reagents 
cause  peptotoxine  to  appear  at  the  expense  of  the 
proteids.  The  evaporation  of  albuminoid  substances 
at  39°  C.  in  presence  of  free  HC1,  leads  to  the  for- 
mation of  bodies  which  by  the  action  of  alcohol  give 
rise  to  peptotoxine.  There  forms,  by  heating  with  a 
feeble  excess  of  the  free  acid,  an  intermediate  body  at 
the  expense  of  the  soluble  albumins.  This  intermedi- 
ate body  (santonine  or  acid  albumin?)  is  decomposed 
by  alcohol,  and  yields  to  the  latter  the  toxic  ptomaine 

*  Salkowsky,  Virchow's  Archiv,  1891,  t.  124,  p  409. 
f  Revue  de  Med.,  1892,  Nos.  i  and  2. 


—  44  — 

which  did  not  exist  before  the  action  of  acid  and  alco- 
hol. These  important  researches  throw  a  new  light 
on  the  question  of  ptomaines. 

The  method  of  Brieger,  employed  in  all  his  re- 
searches, begins  ordinarily  by  the  evaporation  of  the 
first  matters  in  presence  of  hydrochloric  acid,  and 
by  the  extraction  of  the  residue  by  alcohol.  Now 
this  initial  operation  already  gives  rise  to  the  artificial 
production  of  ptomaines.  It  follows  that  the  results 
obtained  by  Brieger  can  give  us  no  information  as  to 
the  poisons  which  pre-exist  in  the  matters  subjected  to 
his  investigation.  They  disclose  to  us  only  the  prod- 
ucts, more  or  less  constant,  of  decomposition  of  these 
matters  as  a  result  of  varied  and  complicated  re- 
actions. And  even,  sometimes,  we  do  not  know 
whence  come  these  products:  from  the  normal  albu- 
mins or  from  microbian  poisons?  Thus,  for  instance, 
although  Kulneff  found,  by  Brieger's  method,  ethyl- 
endiamine  in  the  contents  of  a  dilated  stomach,  we 
believe  with  Bouveret  and  Devic  that  this  body  was 
introduced  by  the  operations. 

What  must  we  think  of  numerous  other  ptomaines 
found  everywhere  by  Brieger  ? 

As  for  the  typhotoxine  of  Brieger,  Bassi  has 
shown  that  it  does  not  pre-exist  in  the  cultures  of  the 
typhoid  bacillus,  but  that  it  may  be  extracted  from 
them  by  the  method  of  Brieger.*  As  it  has  not  been 

*  Bassi:  La  Tifotoxine  di  Brieger  (Gaz.  Chim.  Italiana, 
1889,  t.  18,  p.  521). 


—   45  — 

obtained  by  the  culture  of  any  other  microbe,  it  is 
possible  that  it  may  be  a  product  of  decomposition  of 
the  typhoid  poison. 

In  tetanus,  Kitasato  and  Weyl  have  found  by  the 
method  of  Brieger  the  different  ptomaines  which  he 
had  indicated.  It  is  possible  that  these  ptomaines 
may  be  formed  by  the  decomposition  of  the  primary 
microbian  poison,  and  it  is  certain  that  they  are  not 
the  poison  itself.* 

More  recently,  Baumann  has  found  a  new  method 
for  the  detection  of  diamines.  By  this  method  he 
has  rediscovered  in  cystinuria  many  of  Brieger's 
ptomaines,  and  has  thus  put  beyond  doubt  their  inde- 
pendent existence  in  the  urine.  But  Baumann's 
method,  which  is  based  on  the  employment  of  chloride 
of  benzoyle,  is  too  harsh  for  the  microbian  poisons, 
which  are,  as  we  shall  see  farther  on,  extremely  fra- 
gile.f 

We  see,  then,  that  all  these  researches  on  the 
ptomaines  cannot  give  us  any  idea  as  to  the  chemical 
nature  of  the  microbian  poisons. 

First,  they  depend  on  methods  which  are  too- 
energetic,  which  may  form  artificial  products  even 
with  the  normal  albumins — bodies  relatively  stable. 


*  Kitasato  and  Weyl,  Zeitschrift  f.  Hygiene,  t.  viii,  p. 
404. 

f  Udransky  and  Baumann:  Ueber  das  Vorkommen  von 
Diaminen,  etc.  (Zeitschrift  f.  Physiol.  Chemie,  1889,  *•  13,  p. 
562). 


-  46  - 

And  one  has  no  idea  of  the  decompositions  which 
they  may  effect  in  the  microbian  poisons. 

Secondly,  these  researches  have  been  made  too 
exclusively  from  the  point  of  view  of  pure  chemistry; 
while  the  useful  study  of  the  microbe  poisons  is  im- 
possible without  the  constant  control  of  experimenta- 
tion. Naught  but  experimentation  on  animals  can 
enlighten  us  as  to  the  toxic  effects  proper  to  the 
poisons  of  the  different  microbes.  This  alone  can 
indicate  whether,  after  the  different  chemical  man- 
ipulations, we  have  not  destroyed  or  decomposed  the 
primitive  microbian  poison,  or  substituted  another  for 
it.  This  true  method  of  microbian  toxicology  has 
not  been  followed  by  Brieger  and  other  seekers  after 
ptomaines. 

We  may  note  here  that  Martin,  in  studying  the 
poison  of  charbon,*  comes  to  the  same  conclusion  as 
ourselves;  to  wit,  that  the  ptomaine  which  one  finds 
in  the  anthracoid  cultures  does  not  pre-exist  there, 
but  is  there  combined  with  an  albuminoid  substance. 

On  the  other  side,  microbian  toxicology  has  suc- 
ceeded in  showing,  in  accordance  with  Panum,  that 
the  toxic  products  of  bacteria  are  not  soluble  in 
alcohol. 

Arloing  has  extracted  from  cultures  in  bouillon 
of  the  pneumococcus  liquefaciens  bovis  a  toxic  phlogo- 
genous  substance  which  is  precipitable  by  alcohol, 

*  See  also  in  this  connection,  Seams  Woodhead,  "Bac- 
teria and  their  Products,"  London,  1891. 


—  47  — 

soluble  in  water  and  glycerin,  and  whose  toxicity  is 
destroyed  by  heating  above  110°  C.  Arloing  gave 
the  name  of  diastase  to  this  toxic  substance,  solely 
because  among  the  microbian  products  only  the 
ptomaines  and  diastases  were  known.  The  ^  former 
are  soluble  in  alcohol,  the  latter  are  not.* 

Shortly  afterward,  Christmas  also  observed  that 
the  staphylococcus  aureus  produces  in  cultures  a  basic 
substance  precipitable  by  alcohol. 

Roux  and  Yersin  experimented  with  the  filtrate 
of  diphtheria  cultures  (obtained  through  the  Cham- 
berland  filter),  and  published  several  interesting  re- 
actions of  the  same.  This  poison  is  enfeebled  or  de- 
stroyed by  heating  above  60°  C.  It  is  not  soluble  in 
alcohol.  It  is  carried  down  by  the  different  precipi- 
tates which  form  in  the  midst  of  the  liquid  which 
contains  it;  e.g.,  under  the  action  of  phosphate  of 
lime,  alumina,  etc. 

Roux  and  Yersin  conclude  that  by  all  these  re- 
actions the  diphtheritic  poison  resembles  diastases; 
and  none  the  less  by  its  intense  activity  in  infinitesi- 
mal doses. 

This  identification  of  the  microbian  poisons  with 
the  diastases  was  neither  original  nor  new,  for  it  had 
long  been  a  habit  of  chemists  to  attribute  to  diastases 
every  mysterious  and  unknown  action  exercised  by  a 
substance  of  indeterminate  nature.  Thus  we  have 
seen  that  Virchow,  Schwenninger,  and  Stich  affirmed 

*  Arloing,  "  Les  Virus."     Paris,  1891. 


_  48  — 

the  diastasic  nature  of  the  putrid  poison.  The  ven- 
oms of  serpents  have  for  a  long  time  been  regarded 
as  soluble  ferments.  More  recently  the  poison  of 
jequirity  was  considered  an  enzyme.  This  notion 
has  not,  however,  given  any  real  advancement  to 
science,  for  the  diastases  themselves  are  perfectly 
unknown  to  us  in  their  chemical  properties. 

There  is  one  exception,  however:  the  diastases 
are  characterized  by  their  determinate  chemical 
action  on  other  substances.  Pepsin,  trypsin,  and 
papain  dissolve  albuminoid  substances;  ptyalin  and 
maltine  break  up  starch  into  dextrine  and  glucose; 
invertin  inverts  cane  sugar;  emulsin  and  saponin 
decompose  the  glucosides.  Now  the  only  really 
scientific  way  of  ranking  the  microbian  poisons  among 
the  diastases  would  be  to  identify  them  from  the 
point  of  view  of  this  determinate  chemical  action.  It 
is  necessary  to  show  that  these  poisons  exercise  a 
hydrating  action  on  a  certain  class  of  substances,  and 
that  their  toxic  action  proceeds  precisely  from  this 
chemical  reaction.  No  serious  attempt  has  been 
made  in  this  direction  by  the  partisans  of  the  diastasic 
nature  of  the  microbian  poisons.  They  have  ap- 
pealed, it  is  true,  to  the  great  activity  of  these  poisons 
in  very  small  doses.  But  from  this  point  of  view  we 
should  have  to  rank  among  the  diastases,  hydrocyanic 
acid,  metilcarbylamine,  the  alkaloids  such  as  nicotine 
glucosides  such  as  digitalin,  salts  of  metals  like  cor- 
rosive sublimate,  metalloids  like  phosphorus  and 
fluorine. 


—  49  — 

But  there  are  still  serious  arguments  against  the 
identification  of  the  microbian  poisons  with  the  known 
diastases.  Fermi  and  others  have  studied  the  differ- 
ent enzymes  secreted  by  the  microbes,  and  have  found 
that  they  have  no  appreciable  toxic  action.* 

It  follows  that  the  toxic  action  of  the  bacteria  is 
not  linked  to  their  fermentative  action;  it  does  not 
coincide  with  any  known  diastasic  action.  As  for  any 
new  diastasic  action,  we  cannot  deny  it;  at  the  same 
time  we  cannot  affirm  it  till  we  have  found  it.  It  is 
evident  that  the  toxic  action  of  these  poisons  is  re- 
duced to  some  sort  of  chemical  reaction,  but  we  are 
far  from  knowing  this  reaction.  Consequently,  to 
say  that  the  microbe  poisons  are  diastases,  is  to  speak 
inexactly  if  we  mean  any  diastases  actually  known, 
and  to  juggle  with  words  if  we  suppose  a  new  diastase 
with  unknown  action.  On  the  other  hand,  we  cannot 
deny  the  numerous  analogies  between  certain  poisons 
and  the  soluble  ferments — analogies  which  we  shall 
indicate  later  on.  We  must  come  down  to  the  re- 
searches of  Brieger  and  Frankel  before  we  find  the 
first  serious  attempt  to  determine  the  chemical  nature 
of  the  microbian  poisons.  This  attempt  has  had 
much  success,  although  it  was  not  made  under  the 
conditions  one  could  have  desired. f 


*  Fermi:  Die  Hydrolytische  Enzyme  (Cent.  f.  Physiol- 
ogic, 1891). 

f  Brieger  and  Frankel:  Untersuchungen  tiber  Bacterien- 
gifte  (Berlin.  Klin.  Woch.,  1890,  Nos.  n  and  12). 


4  OOO 


—  5°  — 

Brieger  and  Frankel  first  studied  the  diphtheritic 
poison.  They  found  that  this  poison  behaves  with 
the  different  reagents  like  an  albuminoid  substance. 
This  poison  is  precipitated  by  the  neutral  salts  in  ex- 
cess, such  as  the  sulphate  of  ammonia  and  sulphate 
of  soda.  It  is  not  precipitated  by  sulphate  of  mag- 
nesia. It  is  precipitated  by  absolute  alcohol,  and  is 
very  soluble  in  water.  By  repeated  precipitations  by 
alcohol,  and  dialysis  (contrarily  to  Roux  and  Yersin, 
Brieger  and  Frankel  have  seen  that  the  poison  does 
not  pass  through  the  dialyzing  membrane),  the  sub- 
stance was  obtained  in  a  state  of  purity.  This  sub- 
stance gave  the  following  reactions:  It  is  not  pre- 
cipitated by  ebullition,  sulphate  of  soda,  common 
salt,  sulphate  of  magnesia,  dilute  nitric  acid  (even 
with  the  aid  of  heat),  or  by  acetate  of  lead.  It  is 
precipitated  by  carbonic  acid  (in  saturated  solutions), 
by  the  concentrated  acids,  by  acetic  acid  and  ferro- 
cyanide  of  potassium,  by  phenol,  by  the  organic  acids 
(soluble  in  excess),  by  sulphate  of  copper,  by  nitrate 
of  silver,  by  corrosive  sublimate.  It  is  laevogyrous. 
From  all  these,  and  other  reactions  not  mentioned,  the 
writers  conclude  that  their  toxic  substance  is  exceed- 
ingly like  serum-albumin.  They  have  even  analyzed 
this  substance  and  found  that  it  gives  the  elementary 
composition  of  the  albumins. 

~  Brieger  and  Frankel  have  also  studied,  but  more 
superficially,  the  other  microbe  poisons.  They  have 
found,  beside  the  toxic  albumins,  certain  globulines, 


which  are  distinguished  from  albumins  by  their  in- 
solubility in  distilled  water. 


CHAPTER  VI. 

THE    CHEMICAL    NATURE    OF   THE    BACTERIAL 
POISONS— (Continued). 


SUMMARY. — Criticism  of  the  Work  of  Brieger  and 
Frdnkel — Researches  of  Proskauer  and  Wasser- 
mann — The  Microbe  Poisons  have  Not  yet  been  Pre- 
pared in  a  State  of  Purity — Similar  Non-Microbian 
Poisons —  The  Work  of  Stillmarck — Demonstration 
of  the  Albuminoid  Nature  of  the  Diphtheritic  Poison 
—  Toxicohgical  Differentiation  of  the  Bacterial  Poi- 
sons— Natural  and  Artificial  Poisons:  Their  Char- 
acters—  The  Chemical  Vaccines — The  Antitoxines  or 
Curative  Substances — Hypothesis  as  to  the  Chemical 
Nature  of  the  Poisons — The  Nucleo-albumins  and 
the  Nucleints —  The  Consequences  of  this  Hypothesis. 

We  must  dwell  longer  on  the  toxalbumins  of 
Brieger  and  Frankel,  for  the  researches  of  these 
writers  have  captivated  the  scientific  world  by  their 
apparent  chemical  rigorousness,  and  were  followed 
by  the  creation  of  toxalbumins,  toxalbumoses,  and 
toxopeptones  from  every  source.  The  conclusions  of 
Brieger  and  Frankel,  nevertheless,  call  forth  grave  ob- 
jections. And  first,  in  order  to  prove  that  the  diphther- 
itic bacilli  produce  a  special  albumin,  have  they  elimi- 
nated the  albuminoid  substances  from  the  bouillon  ?• 


—  53  — 

Far  from  that,  they  have  even  added  to  the  ordinary 
bouillon  which  serves  for  the  cultures  the  serum  of 
blood.  How,  then,  can  they  believe  that  this  serum- 
albumin  which  they  find  again  at  the  end  of  their 
purifications  is  not  the  substance  they  introduced, 
carrying  down  in  its  precipitations,  as  Panum  had 
already  shown,  the  true  microbian  poison  ? 

This  supposition  becomes  a  certainty  when  we 
regard  the  feeble  toxicity  of  the  pure  substance  of 
Brieger  and  Frankel,  compared  with  the  far  greater 
toxicity  of  Roux  and  Yersin's  impure  product  of  their 
calcic  precipitate.  In  fact,  Brieger  and  Frankel  have 
less  right  to  say  that  the  diphtheritic  poison  is  a  tox- 
atbumin  than  one  would  have  to  affirm  that  it  is  phos- 
phate of  lime. 

But  phosphate  of  lime  is  a  body  well  known,  and 
it  is  very  easy  to  tell  whether  it  contains  impurities. 
The  albumins  are  much  more  difficult  to  analyze. 
The  proof  of  this  is  that  Brieger  and  Frankel  have 
not  been  able  to  show  us  isolated  the  albumins  of 
broth  and  of  serum,  on  the  one  hand,  and  the  pretended 
toxic  albumin  on  the  other.  There  is  no  doubt,  then, 
that  in  the  experiments  of  Brieger  and  Frankel  the 
diphtheritic  poison  was  always  mixed  with  the  albu- 
mins of  the  culture  media,  and  that  the  chemical 
reactions  described  by  them  were  those  of  the  latter. 

It  may  seem  strange  that  these  writers,  when 
studying  the  chemical  nature  of  the  microbian  poi- 
sons and  finding  them  so  closely  associated  with 


—  54  — 

albumins,  had  not  thought  to  first  separate  the  albu- 
mins from  their  cultures,  leaving  the  microbes  to 
vegetate  in  media  deprived  of  albuminoid  substances. 
But  they  had  the  preconceived  idea  that  the  bacterial 
poisons  could  only  be  formed  at  the  expense  of  the 
albumins  of  the  culture  media.  They  give  us  no 
proof  of  this  notion,  so  widespread  and  yet  so  inexact, 
as  we  shall  show  later  on. 

But  even  apart  from  the  cultures  in  media  de- 
prived of  albumins,  there  were  many  other  means  for 
determining  the  chemical  nature  of  the  bacterial 
poisons. 

We  might,  for  example,  study  what  chemical 
changes  coincide  with  the  heating  of  the  filtered  cul- 
tures above  60°  C. — a  temperature  which  destroys 
the  diphtheritic  poison.  We  offer  this  suggestion 
because  Brieger  and  Frankel  have  made  researches 
which  may  seem  of  a  like  nature.  They  have  studied 
the  non-toxic  albumin  produced  by  the  emasculated 
diphtheria-bacillus,  and  found  it  distinguished  by  sev- 
eral constant  characters  from  the  albumin  containing 
the  diphtheria  poison.  First,  it  is  brown,  while  the 
latter  is  white.  It  is  soluble  in  dilute  alcohol,  and 
combines  with  phenilhydrosine,  while  the  toxic  albu- 
min does  not  combine  with  that  body,  but  with  chlor- 
ide of  benzoyl.  It  contains  more  carbon,  less  oxygen 
and  nitrogen,  than  the  toxic  albumin.  As  the  virulent 
bacilli  are  distinguished  from  the  attenuated  by  the 
energy  of  their  assimilation,  it  is  clear  that  this  dis- 


—  55  — 

tinction  between  the  different  albumins  of  their  cul- 
tures found  by  Brieger  and  Frankel  must  be  of  com- 
plex origin,  and  does  not  permit  us  to  judge  as  to  the 
nature  of  the  diphtheritic  poison., 

We  must  conclude  that,  despite  all  the  chemical 
parade  and  apparatus  of  their  work,  Brieger  and 
Frankel  have  not  succeeded  in  proving  the  legitimacy 
of  their  conclusions  that  the  microbian  poisons  are 
albumins.  Moreover,  the  same  verdict  has  been 
reached  by  the  authors  of  another  work,  emanating 
also  from  Koch's  laboratory — Wassermann  and  Pros- 
kauer. 

These  writers  have  found  in  diphtheritic  cultures 
the  two  albumins  of  Brieger  and  Frankel — the  yellow 
and  the  white.  They  have  seen  that  the  yellow  albu- 
min is  especially  preponderant  in  the  cultures  deprived 
of  toxicity;  it  is  not  itself  toxic.  The  white  albumin 
was  more  abundant  in  the  toxic  cultures,  and  pos- 
sesses a  great  degree  of  toxicity;  by  its  chemical  reac- 
tions this  toxic  albumin  corresponds  to  what  Kuhne 
calls  albumose;  it  is  precipitated  by  alcohol  even  when 
dilute,  by  the  concentrated  acids,  the  salts  of  the 
heavy  metals,  by  potassium  ferrocyanide,  and  by 
acetic  acid.  It  is  not  precipitated  by  ebullition,  by 
nitric  acid,  by  basic  acetate  of  lead,  by  sulphate  and 
chloride  of  sodium  in  excess. 

But  Wassermann  and  Proskauer  are  far  from 
being  prepared  to  identify  the  diphtheritic  poison 
with  this  albumose.  They  remark,  on  the  contrary, 


-  56  - 

that  the  ordinary  bouillon  of  culture  always  contains 
certain  albumoses  by  reason  of  the  infusion  of  meat. 
There  does  not  exist,  therefore,  any  serious  argument 
to  favor  the  view  that  the  toxic  body  which  they  have 
isolated  is  not  the  same  albumose  of  the  bouillon  with 
which  is  found  mingled  the  poison.  This  last  idea  is 
confirmed  by  the  fact  that  different  portions  of  albu- 
mose isolated  from  the  same  toxic  culture  differ  in 
their  toxicity;  they  have,  evidently,  carried  down  vary- 
ing quantities  of  the  poison.  The  German  writers 
conclude  that  the  albuminoid  nature  of  the  diphtheria 
poison  is  possible,  but  that  it  has  not  been  proved. 

From  all  these  researches  we  infer  that  the  diph- 
theria poison  is  closely  associated  with  the  albumin- 
oid substances,  and  more  particularly  with  albumose, 
whose  reactions  it  shares. 

Just  as  in  the  case  of  the  ptomaines,  when  chem- 
ists sought  for  substances  analogous  to  the  alkaloids 
of  vegetal  origin,  the  study  of  microbian  toxalbumins 
clearly  discloses  a  parallel  tendency  in  the  efforts  and 
acquisitions  of  biological  chemistry  and  of  toxicology 
— /'.  <?.,  towards  the  toxic  vegetal  and  animal  proteids. 
In  the  seeds  of  different  plants,  in  the  poisons  of  ser- 
pents, in  the  blood  and  bodies  of  certain  animals, 
were  found  certain  substances  of  extraordinary  viru- 
lence not  belonging  to  the  class  of  alkaloids.  We 
cannot,  unfortunately,  enter  here  into  the  details  re- 
specting all  these  important  researches.  We  will 
content  ourselves  by  mentioning  here  the  writings  of 


—  57  — 

Warden  and  Waddel,  of  Martin  and  of  Kobert,  on 
abrine;  those  of  Weir  Mitchell  and  Edward  Reichart 
and  of  Wolfenden  on  the  poisons  of  serpents;  those 
of  Kobert  on  the  venomous  spiders;  those  of  Mosso 
and  of  Kumahava  on  the  poisonous  fishes,  etc.* 

We  shall  here  analyze  only  one  of  these  memoirs 
— that  of  Stillmarck,  composed  under  the  direction  of 
Kobert,  on  Ricine.  Stillmarck  has  found  that  it  is 
possible  to  extract  from  the  seeds  of  the  ricinus  a 
substance  extraordinarily  toxic,  which  presents  all  the 
reactions  of  albuminoid  bodies.  It  is  not  soluble  in 
alcohol,  in  ether,  or  in  distilled  water.  It  dissolves 
readily  in  saline  solutions.  It  is  precipitated  by  ferro- 
cyanide  of  potassium  and  acetic  acid,  by  saturating 
solutions  containing  it  with  the  neutral  salts,  and  by 
ebullition. 

Stillmarck  supposes  that  this  substance  belongs 
to  the  class  of  albumoses  of  Kuhne.  Unlike  these, 
however,  it  is  coagulated  by  heat. 

As  for  the  important  question  whether  we  can 
identify  the  poison  with  this  albuminoid  substance,  or 
if  the  poison  is  simply  mixed  with  the  latter,  Still- 
marck advances  serious  arguments  in  support  of  the 
first  alternative.  First,  by  boiling,  ricine  is  coagulated 
and  rendered  completely  insoluble;  at  the  same  time, 
its  toxicity  is  completely  abolished.  Then  again,  pan- 


*Vide  the  entire  literature  in  Hulliburton's  Text-Book 
of  Chemical  Physiology  and  Pathology.     London,  1891. 


-  58  - 

creatic  juice,  which  digests  albuminoid  substances, 
destroys  also  the  toxicity  of  ricine. 

It  is  rendered  very  probable  by  these  re- 
searches, that  there  exist  certain  albuminoid  bodies 
of  nature,  yet  undefined,  which  are  very  violent  poi- 
sons. We  have  recently  shown  that  the  diphtheria 
poison  must  also  be  ranked  in  this  category,  for  it  is 
destroyed,  or  at  least  modified,  by  the  ferments  of  the 
albuminoid  bodies. 

Now  to  what  class  of  albuminoid  bodies  do  the 
microbian  poisons  belong?  This  question  is  the  more 
important  because  these  poisons  have  an  evident 
kinship  to  a  large  class  of  vegetable  and  animal  poi- 
sons of  which  we  have  before  spoken. 

Unfortunately,  the  data  which  biological  chem- 
istry possesses  respecting  the  albuminoid  matters  in 
general  are  still  very  incomplete  and  indefinite.  The 
methods  for  their  analysis  are  still  very  imperfect.  It 
is  evident  that  under  these  conditions  one  cannot 
rigorously  determine  the  chemical  nature  of  the 
microbian  poisons.  We  have  already  seen  that  even 
the  best  studied  of  all  these  poisons,  that  of  diph- 
theria, has  not  yet  been  obtained  in  a  state  of  purity,, 
unmixed  with  the  albumins  of  the  culture  media. 

However,  we  possess  for  the  microbian  poisons  a 
precious  and  powerful  means  of  analysis,  namely,  ex- 
perimentation, which  is  based  on  the  property  of  these 
poisons  to  produce  in  animals  certain  phenomena 
similar  to  those  of  the  infectious  diseases:  intoxica- 
tion and  immunity. 


—  59  - 

This  means  has  enabled  us  to  establish  a  funda- 
mental distinction  between  two  classes  of  microbian 
poisons:  the  natural  or  primary  poisons,  and  the 
modified  or  artificial  poisons.  I  have  shown  that  in 
cholera  there  exist  two  poisons  totally  different  in 
their  physiological  effects;  the  one  produces  the  diar- 
rhcea,  while  the  other  has  a  phlogistic  action.  These 
poisons  differ  also  by  their  resistance  to  heat;  the 
first  is  destroyed  by  heating  it  above  60°  C.,  while  the 
second  supports  a  temperature  of  120°  C.  for  several 
hours.  It  is  a  tenable  supposition  that  these  two 
poisons  are  closely  associated  together,  and  that  the 
second  comes  from  the  decomposition  of  the  first.* 

As  for  the  diphtheritic  poison,  I  have  also  found 
that  it  is  necessary  to  distinguish  the  secondary 
poison  which  appears  on  the  decomposition  of  the 
primary  poison  by  heat,  by  acids,  by  the  soluble  fer- 
ments. 

The  attentive  study  of  the  other  different  poisons 
enables  us  to  classify  them  also  in  one  or  the  other 
of  these  two  groups,  which  we^hall  characterize  more 
in  detail. 

The  natural  poisons  correspond  to  those  which 
Brieger  and  Frankel  have  called  the  toxalbumins,  and 
Klemperer  the  toxines.  They  are  substances  which 
reproduce  more  or  less  exactly  the  symptoms  of  the 


*Gamale!a:  Experimental  Researches  on  the  Poisons 
of  Cholera  (Arch,  de  Med.  Experimentale,  1892,  No.  2). 


—  6o  — 

infectious  disease.  They  are  very  unstable,  and  are 
decomposed  by  a  temperature  above  60°  C.  The 
animals  which  are  refractory  to  the  disease  produced 
by  their  microbe,  are  also  refractory  to  the  primary 
poison  of  this  microbe. 

The  modified  poisons,  which  are  also  named 
proteins  (Buchner,  Klemperer),  do  not  reproduce  the 
typical  phenomena  of  the  microbian  disease.  They 
cause  hypothermia  or  fever,  according  to  the  dose, 
inflammation  more  or  less  intense  at  the  point  of  in- 
oculation, and  symptoms  more  or  less  pronounced  of 
cachexia.  They  have  the  remarkable  property  of 
exciting  tuberculous  animals  to  general  and  local  re- 
action. These  poisons  resist  boiling.  They  are  pre- 
cipitated by  alcohol.  Panum's  poison  probably  be- 
longs to  this  class  of  substances.  These  poisons  have 
nothing  to  do  with  the  production  of  immunity.  In 
certains  cases,  on  the  contrary,  they  have  an  opposite 
influence,  predisposing  the  economy  to  the  invasion 
of  microbes.  The  vaccinated  animals  are  not  refrac- 
tory to  these  artificial  poisons.  By  the  side  of  these 
primary  and  secondary  poisons,  experimental  analysis 
has  revealed  still  a  third  and  a  distinct  order  of 
substances.  These  are  the  chemical  vaccines — the 
microbian  products  which  confer  on  animals  immu- 
nity against  infection  by  the  living  microbe.  In 
certain  cases  the  chemical  vaccination  may  be  ob- 
tained by  means  of  the  primary  poison  — as,  for  in- 
stance, in  the  vaccinations  against  the  septic  vibrio, 


—  6i  — 

or  against  the  streptococcus  of  pneumonia.  But  in 
many  other  cases  the  primary  poisons  do  not  confer 
immunity,  but  lead,  even  in  small  doses,  to  cachexia. 
Such  is  the  case  with  regard  to  tetanus  and  diphtheria. 
But  here  also  a  chemical  vaccination  is  possible  by 
means  of  certain  artifices.  Thus,  for  instance,  we 
prepare  the  vaccines  by  heating  the  primary  poisons 
above  their  critical  point,  to  70°  or  80°  C.  We  may 
suppose  that  this  heating  sets  free  the  vaccinal  sub- 
stance of  the  primary  poison.  This  method  succeeds 
for  pneumonia,  and  sometimes  for  tetanus  and  diph- 
theria. In  some  diseases  chemical  vaccination  is  easier; 
the  microbian  products  vaccinate,  even  after  having 
been  subjected  to  a  temperature  of  100°  and  even 
120°  C.  This  is  the  case  with  cholera  and  the  vibri- 
onic  septicaemia,  which  I  was  the  first  to  discover. 

In  certain  cases  the  vaccinant  products  have 
been  quite  isolated  from  the  primary  or  modified 
poisons.  I  have  found  that  against  cholera  and 
against  the  avicide  vibrio  we  may  vaccinate  with  the 
volatile  products  of  the  cultures.  As  for  the  proteus 
vulgaris,  it  has  been  seen  that  we  can  vaccinate  by 
the  different  ptomaines,  such  as  neuridine,  muscarine, 
etc.  Thus  we  see  that  the  chemical  vaccines  are  very 
varied.  We  may  say,  in  general,  that  they  are  more 
stable  than  the  corresponding  poisons.  I  have  re 
garded  them  as  a  decomposition-product  of  the  poi- 
sons; in  any  event,  they  have  a  variable  chemical, 
nature,  being  decomposed  by  ebullition. 


—    62    — 

There  exist,  lastly,  substances  still  more  com- 
plex, which  are  found  only  in  the  organism  of  re- 
fractory animals  after  an  injection  of  the  poisons. 
These  substances,  which  are  combinations  of  poisons 
with  the  animal  substances,  or  possibly  chemical  vac- 
cines quite  pure,  have  the  property  not  only  of  vac- 
cinating the  animals  against  an  infection  to  come, 
but  of  curing  the  disease  when  declared.*  The  nature 
of  these  immunizing  and  curative  substances  is  com- 
pletely unknown. 

If  we  wish  now  to  sum  up  all  this  long  study  of 
the  chemical  nature  of  the  microbian  poisons,  we 
arrive  at  the  following  conclusions: 

Among  the  products  of  the  microbes,  there  are 
four  classes  of  substances:  First,  the  organic  bases 
(which,  if  they  do  not  exist  essentially  as  ptomaines, 
assume  that  form  under  the  different  processes  of 
analysis);  second,  certain  substances  of  albuminoid 
nature,  or  primitive  poisons,  easily  decomposible  at  a 
temperature  above  60°  C.;  third,  still  other  albumin- 
oid substances,  which  are  more  stable — the  modified 
poisons;  fourth,  a  class  of  substances  which  can  only 
be  defined  by  their  physiological  action,  the  chemical 
vaccines — these  are  associated  with  one  of  the  three 
preceding  classes  of  microbian  products. 

Despite  all  this  diversity  of  the  microbian  prod- 
uts,  and  all  the  apparent  complexity  of  their  relations, 


*GamaleIa:  Immunisation  (Gaz.  Hebd.,  1891,  No.  47). 


-63  - 

we  may  frame  a  hypothesis  to  explain  their  chemical 
nature. 

Biological  chemistry  has  taught  us  that  there  is  a 
particular  class  of  substances  more  complicated  than 
the  ordinary  albuminoid  substances,  extraordinarily 
unstable,  decomposing  even  by  prolonged  contact 
with  alcohol  and  by  heat  above  60°  C.,  giving  rise  by 
their  decomposition  to  the  formation  of  other  albu- 
minoid substances  more  stable,  as  well  as  to  the  dif- 
ferent ptomaines  and  leucomaines.  These  are  the 
nucleo-albumins  or  vitellines,  constituent  parts  of  all 
animal  and  vegetable  cells.  I  have  made  the  hypo- 
thesis that  the  natural  poisons  of  the  bacteria  are 
precisely  such  nucleo-albumins,  which  give  rise  to 
nucleines  or  modified  poisons  by  their  decomposition, 
and  to  ptomaines  by  a  more  profound  decomposition. 
The  principal  chemical  character  of  the  nucleo-albu- 
mins is  the  great  quantity  of  phosphorus  which  they 
contain.  In  the  microbian  poisons,  chemists  have 
already  found  much  phosphorus.  Such  is  the  case  in 
regard  to  the  poisons  of  cholera  (Petri),  the  avicide 
vibrio  (Wolkow),  and  Koch's  tuberculin.  But  all  these 
acquisitions  of  organic  chemistry  have  no  decisive 
value,  for  no  one  of  these  poisons  has  been  obtained 
in  a  state  of  purity — all  have  been  mixed  with  other 
substances  derived  from  the  culture  media.  Till  the 
determination  of  phosphorus  shall  have  been  made 
for  a  perfectly  pure  microbian  poison,  our  idea  will 
remain  only  an  hypothesis.  But  it  may  be  verified 


—  64  — 

upon  other  characters  of  the  nucleo-albumins — char- 
acters which  are  less  decisive  from  a  chemical  point 
of  view,  but  much  more  interesting  from  the  stand- 
point of  microbian  toxicology. 

And,  first,  there  arises  the  fundamental  question: 
Are  the  microbian  poisons  the  products  of  decom- 
position of  the  nutritive  media  of  the  bacteria,  as  cur- 
rent opinion  has  it,  or  are  they  the  constituent  parts- 
of  the  bodies  of  the  microbes,  as  they  ought  to  be  if 
they  were  nucleo-albumins?  This  important  question 
concerning  the  origin  of  the  microbian  poisons  wilt 
be  examined  in  the  next  chapter. 


CHAPTER  VII. 

THE    ORIGIN    OF    THE    MICROBIAN     POISONS. 


SUMMARY. — Preconceived  Idea  that  the  Poisons  Can 
Only  Come  from  the  Decomposition  of  Albumin- 
oid Bodies — Experiments  Already  Old  which  Over- 
throw this  Idea — Researches  of  Polotebnoff,  Popoff, 
Bergmann,  Schuller — Numerous  Researches  Made 
at  the  Laboratory  of  Pachoutine — Recent  Labor  of 
Guinochet — The  Microbian  Poisons  Are  Not  Prod- 
ucts of  Decomposition,  but  the  Result  of  Synthesis — 
Are  They  Secretions  ? — Arguments  in  Favor  of  this 
Idea — Arguments  which  Combat  It — The  Poisons 
are  Intimately  Linked  to  the  Bodies  of  the  Bacteria 

—  The  Experiments  of  Cantani  and  of  the  Author 

—  The  Researches  of  Buchner  on  the  Proteins — 
Criticism   of  these    Researches — All    the    Microbe 
Poisons  Come  from  the  Bodies  of  Bacteria  —  Utility 
of  this  Source  for  the  Microbes. 

In  all  the  history  of  the  researches  on  the 
microbian  poisons,  the  idea  was  generally  admitted 
that  these  poisons  take  their  origin  in  the  course  of 
the  decomposition  of  albuminoid  matters.  This  idea 
is  prevalent  at  the  present  time.  Nevertheless,  there 

exist  in  science  many  facts  which  completely  over- 

5  ooo 


—  66  — 

throw  it.  As  early  as  the  epoch  of  the  study  of  the 
putrid  poison,  chemists  were  seeking  to  eliminate  the 
albuminoid  substances  from  the  matters  undergoing 
putrefaction,  in  order  to  render  easier  the  extraction 
of  the  putrid  poison.  Thus,  for  instance,  Bergmann 
and  Schmiedeberg  had  employed  brewer's  yeast  in 
their  famous  experiments  on  sepsine. 

Polotebnoff  was  the  first  to  employ  an  entirely 
mineral  medium  and  allow  it  to  putrefy.  It  was 
Pasteur's  liquid,  composed  of  ammonia  and  of  dif- 
ferent neutral  salts.  Polotebnoff  found  that  the 
liquid  when  putrefied  is  not  more  toxic  than  the  fresh 
liquid. — Popoff,  who  repeated  the  experiments  of 
Polotebnoff,  has  shown  that  these  conclusions  are 
not  exact,  and  that  Pasteur's  liquid  when  putrefied 
produces  absolutely  the  same  symptoms  of  intoxica- 
tion as  the  putrefied  albuminoid  matters.  But  Popoff 
did  not  kill  the  microbes  in  his  liquid,  and  he  sup- 
posed that  it  was  precisely  the  microbes  which  pro- 
voked this  septic  intoxication. — The  same  experiments 
were  made  by  Hugo  von  Brehm. — Bergmann  has 
given  to  the  question  a  decisive  advancement.  He 
has  shown  that  Pasteur's' liquid  when  putrefied  pre- 
serves all  its  septic  action,  even  after  having  been 
boiled. — Anders  has  also  found  that  neither  boiling 
nor  filtration  through  clay  destroys  the  septic  action 
of  this  liquid. 

Thus  was  established  this  principle  of  the  highest 
importance:  That  the  septic  poison  is  the  result  of 


-  67  - 

the  synthetical  action  of  the  microbes,  and  not  of  the 
decomposition  of  the  putrid  matters. 

The  same  results  were  obtained  by  Schuller,  who 
operated  with  the  liquid  of  F.  Cohn,  which  contains 
no  sugar,  and  whose  sole  organic  compound  is  am- 
monium tartrate. 

These  experiments  have  since  been  often  repeat- 
ed with  the  same  positive  results.  These  researches 
have  been  chiefly  made  at  the  laboratory  of  Pachou- 
tine.  We  must  mention  especially  the  theses  of  Bott- 
charoff  and  of  Cosorotoff  emanating  from  this  labora- 
tory. All  these  writers,  as  well  as  numerous  others 
who  have  studied  the  details  of  putrid  intoxication — as, 
for  instance,  the  fever  and  the  gaseous  exchanges- 
have  uniformly  arrived  at  this  result:  that  the  putrid 
poison  is  formed  in  the  absence  of  every  albuminoid 
body,  and  that  it  is,  consequently,  the  result  of  the 
synthetic  activity  of  the  microbes.  Note  also  that 
these  writers  have  .confirmed  in  general  the  datum 
of  Panum  as  to  the  great  toxicity  of  the  products 
precipitated  by  alcohol.  Nevertheless,  contrarily  to 
Panum,  the  alcoholic  extract  had  often  itself  also  a 
certain  septic  activity. 

Unfortunately,  all  these  experimental  researches 
were  carried  on,  not  with  pure  cultures  of  bacteria, 
but  with  cultures  containing  the  microbes  of  putre- 
faction in  indefinite  quantity.  They  were  carried  on 
from  the  point  of  view  of  experimental  pathology, 
and  bacteriology,  which  has  mostly  confined  itself  to 


—  68  — 

pure  cultures  of  microbes,  has  rather  ignored  the  im- 
portant results  established  by  these  labors. 

Of  much  more  importance  are  the  researches  of 
Guinochet  on  the  diphtheritic  poison.  Guinochet  has 
found  that  the  diphtheritic  bacillus,  which  is  easily 
cultivated  in  normal  filtered  human  urine  containing 
not  a  trace  of  albumin,  produces  its  poison  in  that 
medium  quite  as  energetically  as  in  meat  broth.* 

Thus  we  find  that  there  is  little  ground  for  the  pre- 
conceived idea,  which  has  lasted  so  long,  that  the 
pathogenic  microbes  fabricate  their  poisons  by  a 
special  decomposition  of  albuminoid  matters.  This 
notion,  demonstrated  false,  must  give  place  to  the 
more  just  conception,  viz.,  that  the  poisons  are  the 
synthetic  products  of  the  microbes. 

Independently  of  the  theoretical  importance  of 
this  conclusion  of  Guinochet,  we  must  also  indicate 
its  practical  side.  We  shall  be  able  henceforth  to 
study  the  formation  of  the  micrabian  poisons  in  con- 
ditions more  simple  than  those  which  were  chosen 
by  Brieger  and  Frankel  and  their  successors,  in  a 
medium  totally  deprived  of  albumins,  f 

The  question  now  presents  itself:  What  is  the 
role  of  these  substances  ?  What  place  do  they  oc- 


*  Guinochet:  Labors  of  the  Laboratory  of  Straus,  at  the 
Ecole  de  Medecine,  Paris,  France. 

fit  only  remains  to  find  a  culture  medium  favorable  to 
the  development  of  the  microbes,  for  the  mineral  media 
(liquids  of  Pasteur,  Noegeli,  and  Cohn)  are  not  suitable  for 


-69- 

cupy  in  the  life  of  the  microbes  ?  For  the  definition 
of  animal  poisons  depends  for  its  significance  upon 
the  relation  of  these  poisons  to  the  higher  organisms, 
and  not  to  microbe-biology. 

There  were  already  a  certain  number  of  syn- 
thetic and  specific  products  known  to  microbiology, 
whose  signification  was  sufficiently  clear.  These 
were  the  soluble  ferments,  or.  diastases,  which  the 
microbes  secrete  to  render  assimilable  the  aliments 
which  surround  them.  The  yeast-plant  secretes  inver- 
tine,  which  decomposes  cane  sugar,  not  directly  utiliz- 
able,  into  dextrine  and  levulose.  The  bacillus  of  lactic 
acid  secretes  the  lab-ferment  (which  serves  to  coagu- 
late the  casein  of  milk),  and  casease  (which  dissolves 
and  peptonizes  it). 

It  was  natural  to  think  of  the  similarity  of  these 
microbian  manifestations  to  the  empoisonments  pro- 
duced by  the  pathogenic  microbes.  One  might  readily 
believe  that  the  poisonous  secretions  serve  also  to 
prepare  the  animal  soil  for  the  microbian  life,  whether 
by  rendering  the  animal  humors  assimilable  for  the 
microbes,  or  by  abolishing  the  vital  resistance  of  the 

the  pathogenic  bacteria.  I  have  found  that  they  grow  well 
in  the  following  medium: 

Water 1000 

Liebig's  extract 5 

Glycerin 4° 

Common  salt 5 

Liebig's  extract  is  totally  exempt  from  albuminoid 
matters. 


—  70  — 

animals  and  thus  rendering  the  field  clear  for  the  oc- 
cupancy of  the  microbes. 

This  notion  soon  found  a  certain  support  in 
experimentation.  It  has  been  observed  that  the 
diphtheritic  and  tetanus  poisons  behave  towards  the 
different  reagents  precisely  like  the  soluble  ferments. 
It  has  also  been  noted  that  different  substances — the 
soluble  ferments  of  papain;  such  microbian  products 
as  the  poison  of  the  bacillus  prodigiosus;  the  poisons 
of  plants,  like  abrine — also  possess  the  property  of 
rendering  the  animals  in  which  we  inject  them  in  min- 
imum doses  predisposed  to  the  pullulation  of  microbes 
which  are  not  pathogenic  to  the  same  animals  in  the 
normal  state.* 

Nevertheless  this  comparison  of  the  poisons  to 
the  secretions  accounts  but  superficially  for  the  facts. 

First,  there  exist  certain  pathogenic  bacteria 
which  produce  such  diseases  (of  the  highest  import- 
ance) as  tuberculosis,  cholera,  and  charbon,  where 
we  do  not  find  any  toxic  secretions. 

Then  again,  even  the  production  of  poisons  in 
diphtheria  and  tetanus  has  a  very  superficial  corre- 
spondence to  a  secretion.  In  fact,  the  quantity  of 
these  poisons  in  the  culture  liquids  does  not  grow 
proportionately  -to  the  development  of  the  bacteria 
themselves.  On  the  contrary,  an  attentive  observa- 
tion reveals  the  following  facts: 

*See  GamaleJa  "On  the  Reproduction  of  Cholera  in 
the  Hare"  (Congress  of  Berlin,  1890). 


At  the  very  first,  and  at  the  most  active  moment 
of  the  life  of  the  microbes,  the  liquid  of  culture  is  acid 
and  devoid  of  all  toxic  power.  Later,  when  the  bac- 
teria have  ceased  to  multiply  and  are  deposited  at 
the  bottom  of  the  culture-flask,  the  liquid  becomes 
alkaline  and  more  and  more  toxic.  Its  toxicity  aug- 
ments, in  a  certain  measure,  progressively  with  the 
duration  of  the  sojourn  of  the  bacteria  in  their  alka- 
line liquid  of  culture. 

The  explanation  of  this  fact  is  very  simple.  The 
poison  is  contained  inside  the  bodies  of  the  bacteria, 
-and  is  extracted  but  slowly  by  the  alkaline  liquid  in 
which  they  are  macerated. 

This  hypothesis  as  to  the  origin  of  all  the 
microbe  poisons  from  the  bodies  of  the  bacteria  has 
been  already  proposed  by  Cantani  a  propos  of  the 
cholera  poison.  Cantani  supposed  that  in  the  cholera 
cultures  it  was  the  bacilli  themselves  which  were 
toxic,  like  certain  fungi  which  cause  poisoning  when 
injected.  He  did  not  prove  his  hypothesis,  which  long 
remained  without  support.  This  was  six  years  ago, 
.and  Cantani's  article  "Die  Giftigkeit  der  Cholera- 
bacillen  "  was  published  in  the  Deutsche  Medicinische 
Wochenschrift  in  1886.  Since  then  I  have  shown 
that  the  cadavers  of  divers  bacteria  remain  very 
toxic  even  after  having  been  exposed  to  a  heat  of 
100°  to  120°  C. 

Later,  I  showed  that  we  can  prepare  from  the 
cadavers  of  the  cholera  vibrio  and  of  the  avicide  vibrio 


—  72  — 

very  toxic  extracts  which  possess  at  the  same  same 
vaccinal  properties.* 

At  the  same  time  that  this  study  of  the  general 
action  of  the  bacterial  cadavers  was  being  pursued, 
other  experimenters  were  investigating  their  local 
effects.  After  Gravitz  and  Wyssokowitch  had  shown 
that  the  cadavers  of  certain  bacteria  have  the  prop- 
erty of  producing  suppuration  in  animals,  Buchner 
made  of  this  question  a  special  study.  By  boiling 
with  dilute  alkalies,  he  succeeded  in  extracting  from 
the  bodies  of  different  bacteria,  such  as  the  typhoid 
and  the  pyocyanic  bacillus,  substances  in  general  of 
little  toxicity,  but  possessing  the  property  of  producing^ 
by  subcutaneous  injection  an  exudative  inflamma- 
tion. With  doses  still  more  feeble,  these  poisons 
provoke  only  an  emigration  of  leucocytes  (positive 
chimiotaxy). 

By  their  chemical  properties  these  substances 
correspond  quite  well  to  those  which  Rencki  had 
described  under  the  names  of  myco-protein  and  antraco- 
protein.  Buchner  considered  them  as  alkali-albumins 
or  proteins,  and  under  that  name  they  have  since 
been  studied.  Their  most  curious  property  has  been 
set  forth  by  Koch.  Already  in  1889  I  had  indicated 
that  tuberculous  guinea-pigs  have  an  extreme  sus- 
ceptibility towards  the  vibrio  poison.  Koch  has  sue- 

*GamaleIa,  in  Ann.  de  1"  Instit.  Pasteur,  1888,  No.  5; 
also  1889,  No.  10;  also  in  Compt.  Rend,  de  la  Soc.  de 
Biologic,  Nov.  30,  1890. 


T  3 
/  O 

ceeded  in  extracting  from  the  tubercle  bacilli,  by 
heating  with  glycerin,  a  substance  called  tuberculin, 
which  proves  to  be  extremely  active  in  tuberculous 
subjects.  It  provokes  in  them  the  phenomena,  so 
often  described,  of  general  and  local  reaction.  This 
reaction  of  the  tuberculous  to  tuberculin  has  long 
been  wrongly  considered  as  a  characteristic  effect 
of  the  latter.  But  latterly  it  has  been  shown  that, 
besides  the  avicide  vibrio  and  the  bacillus  tuber- 
culosus,  many  other  bacteria  contain  a  substance  hav- 
ing a  similar  property.  Buchner  and  Rohmer  have 
given  the  following  directions  for  the  extraction  of 
these  substances:  Dry  the  bacteria  after  separating 
them  from  their  culture  media;  boil  them  several 
hours  in  alkaline  water,  and  leave  the  decoction  to 
macerate  for  several  days  at  the  temperature  of  37° 
C;  then  filter  the  liquid,  and  precipitate  the  active 
substance  by  acetic  acid. — Thus  we  see  that,  with  the 
proteins  of  Buchner,  the  poisons  extracted  from  the 
bodies  of  bacteria  have  acquired  the  right  of  place  in 
bacteriology.  But  we  must  make  some  important 
reservations  in  regard  to  this  conception  of  the  pro- 
teins. 

First,  Buchner  and  the  chemists  who  followed 
him  established  a  fundamental  distinction  between 
the  proteins  and  the  toxalbumins  or  toxines — for  the 
latter  come  from  the  secretions  of  the  bacteria,  the 
former  only  from  their  bodies.  The  proteins  exercise 
a  positive  chimiotaxy,  the  toxines  negative  chimio- 


—   74  — 

taxy.  (We  shall  return  shortly  to  the  role  of  micro- 
bian  poisons  in  inflammation.)  As  to  the  source  of 
these  poisons,  we  have  already  seen  that  there  are 
reasons  to  suppose  that  the  toxalbumins  are  also  con- 
tained in  the  bacteria.  If  Buchner  has  not  succeeded 
in  extracting  from  the  bodies  of  microbes  aught  but 
the  proteins,  this  is  evidently  due  to  his  crude  method 
of  decoction,  which  surely  destroys  the  toxalbumins. 
We  have  shown  that  in  cholera  one  may  obtain  by 
extraction  at  a  temperature  of  55°  to  60°  C.  another 
poison  than  that  which  is  extracted  by  ebullition. 
Likewise,  in  respect  to  tuberculosis,  if  instead  of 
Koch's  process  you  employ  Weyl's  more  efficacious 
method  of  extraction  by  alkaline  water  at  low  tern- 
peratures,  you  obtain  a  poison  more  active  than 
tuberculin,  and  endowed  with  other  physiological  and 
chemical  properties.  Now  it  is  more  simple  to  sup- 
pose that  all  the  poisons  of  the  microbes  come  from 
their  bodies;  but  that  the  extraction  at  low  tempera- 
tures gives  us  the  toxalbumins  or  primary  poisons,  and 
that  ebullition  gives  the  proteins  or  modified  poisons. 
Moreover,  in  coming  back  to  the  point  of  de- 
parture of  the  researches  of  Buchner — to  the  suppura- 
tion produced  by  the  microbian  cadavers — it  is  proper 
to  note  that  this  savant  has  completely  failed  in  his 
attempt  to  find  the  pyogenic  substances.  The  pro- 
teins which  he  has  isolated  produce  an  exudative 
inflammation,  and  not  suppuration.  Buchner  has 
found  the  likeness  of  suppuration  in  the  emigration 


—  75  — 

-of  the  leucocytes,  in  the  positive  chimiotaxy  provoked 
by  his  proteins.  But  this  positive  chimiotaxy  not  only 
has  not  the  significance  of  suppuration,  but  is 
not  even  inflammation.  It  is  nothing  but  hyper- 
semia.  We  see  it  when  we  introduce  very  small  doses 
of  proteins  (in  capillary  tubes)  under  the  skin  of 
animals.  When  we  augment  the  doses  injected,  we 
observe  (as  we  have  seen)  the  production  of  a  serous 
exudative  inflammation  without  leucocytes.  So  our 
positive  chimiotaxy  becomes,  by  the  augmentation  of 
the  doses,  negative  chimiotaxy,  and  cannot  serve  to 
distinguish  the  two  classes  of  poisons.  But  suppura- 
tion is  not  obtained  with  the  proteins. 

It  is  very  probable  that  the  failure  of  Buchner 
to  extract  pyogenic  substances  from  the  microbian 
bodies  was  due  to  his  faulty  process  of  extraction, 
and  that  the  pyogenic  substances  belong  to  the  cate- 
gory of  the  more  fragile  (the  primary)  poisons.  As  to 
the  chemical  nature  of  the  substances  extracted  by 
Buchner  from  the  bodies  of  the  bacilli,  their  identity 
with  the  class  of  alkali-albumins  or  proteins  cannot 
longer  be  admitted.  Buchner  had  found  that  these 
substances  present  the  reaction  of  albuminoids,  and 
that  they  are,  besides,  precipitated  by  neutralizing  with 
acetic  acid  their  alkaline  solutions.  But  this  latter  re- 
action is  an  artificial  one,  produced  by  reason  of  their 
extraction  by  alkalies  from  the  bodies  of  bacilli.  When 
Buchner  later  employed  for  this  extraction  neutral 
water,  he  no  longer  observed  precipitation  by  the 


—  76  — 

acids.  If  we  still  continue  to  call  these  substances 
alkali-albumins,  it  is  by  an  abuse  of  language.  Like- 
wise, Koch's  tuberculin,  which  belongs  by  all  its 
physiological  properties  to  the  same  class  as  Buchner's 
proteins,  but  which  is  extracted  by  means  of  a  neutral 
solvent,  does  not  present  the  characteristic  reactions 
of  the  alkali-albumins.  It  is  evident,  consequently, 
that  the  substances  of  Buchner,  or  our  modified 
poisons,  are  not  alkali-albumins.  We  have  already 
stated  in  the  preceding  chapter  that  we  believe  them 
to  be  nucleines.  As  for  tuberculin  especially,  our 
hypothesis  is  found  confirmed  by  the  richness  of  this 
substance  in  phosphorus,  which  we  find  even  after 
having  freed  from  phosphoric  salts  the  bouillon  in 
which  the  bacillus  tuberculosis  has  vegetated. 

Thus  the  modern  researches  on  the  origin  of  mi- 
crobian  poisons  lead  us  to  the  same  results  as  the  study 
of  their  chemical  properties.  The  microbian  poisons 
may  be  divided  into  primary  and  secondary.  They 
are  the  results  of  the  creative  synthesis  of  the 
bacteria,  and  come  from  their  bodies.  Thus  is  again 
confirmed  our  hypothesis  respecting  their  chemical 
nature  which  identifies  them  with  the  constituent 
parts  of  the  nuclei  of  cells— the  vitellines  and  the 
nucleines. 

The  toxicity  of  the  microbian  bodies  may  have 
also  a  teleological  signification,  explicable  from  the 
point  of  view  of  the  struggle  for  existence  and  of 
natural  selection;  for  the  toxicity  of  the  bodies  of 


—  77  — 

bacteria  may  be  useful  to  them  as  a  means  of  pre- 
serving them  against  aggressors  which  would  devour 
them. 


CHAPTER  VIII. 

ACTION  OF  THE  BACTERIAL  POISONS  ON  THE 

ANIMAL  ORGANISM— ACCUSTOMANCE 

AND  IMMUNITY. 

SUMMARY.  — Insufficiency  of  our  Knowledge  of  the  Mode 
of  Action  of  the  Bacterial  Poisons  on  the  Animal 
Organism — Local  Action  of  the  Modified  Poisons — 
Selective  Action  of  Tuberculin  and  Malleine — De- 
struction of  Poisons  in  the  Bodies  of  Refractory 
Animals — Explanations  of  the  Different  Contradic- 
tions— Re'sume'  and  Conclusions  on  the  General 
Toxicity  of  Microbes. 

As  the  microbian  poisons  have  not  been  the 
object  of  systematic  researches  in  their  aggregate,  cer- 
tain important  questions  concerning  their  history  are 
still  undecided.  The  mode  of  action  of  the  poisons 
on  the  organism  is  one  of  these  questions.  Even  in 
the  case  of  the  best  known  of  the  microbian  poisons, 
we  do  not  yet  know  on  what  elements  they  act  pref- 
erably, whether  on  the  nerve-cells  or  muscles,  on 
the  heart  or  capillary  vessels,  or  even  on  the  blood. 

Most  of  these  poisons  produce  a  local  lesion  at 
the  place  of  injection.  This  local  lesion  may  go  on 
from  a  serous  or  sero-sanguinolent  exudation  to 
necrosis.  This  phlogistic  action  belongs  to  the- 


—  79  — 

majority  of  the  modified  poisons.  It  is,  too,  a  little 
singular  that  when  injected  into  the  blood,  these  same 
poisons  have  a  marked  antiphlogistic  action.  The 
intravenous  injection  of  these  poisons  prevents  the 
appearance  of  inflammation,  even  when  solicited  by 
very  energetic  agents,  such  as  the  application  of  cro- 
ton  oil.  Bouchard  has  endeavored  to  explain  this 
antiphlogistic  action  by  the  paralysis  of  the  vaso- 
motor  centres;  but  this  explanation  is  not  fully 
demonstrated,  and  still  remains  contested  (Samuel).* 

When  injected  under  the  skin,  but  in  very  small 
doses,  the  same  modified  poisons  provoke  inflamma- 
tion, not  at  the  place  of  their  introduction,  but  around 
foci  of  pre-existent  lesions.  The  best  known  examples 
of  this  elective  action  are  the  local  reactions  provoked 
by  tuberculinf  in  the  tuberculous,  and  by  malleinej  in 
horses  affected  with  glanders. 

The  attempt  has  seriously  been  made  to  explain 
this  elective  inflammation  by  the  positive  chimiotaxy 


*Charrin  and  Gamaleia :  On  Inflammation  (C.  R.  de 
la  Societ6  de  Biologic,  July  5,  1890).  Gamaleia  :  On  the 
Local  Lesion  in  Microbian  Diseases  (Arch,  de  Med.  Exper., 
1891,  No.  2). 

f  Koch,  Deutsche  Medicinische  Wochen.,  1890,  No.  46. 
Gamaleia:  On  the  Treatment  of  Tuberculosis  by  Koch's 
Method  (Arch,  de  Med.  Experimentale,  1891,  No.  2). 

JHelmann:  On  the  Clinical  and  Experimental  Diag- 
nosis of  Glanders  (Messager  de  la  Science  Veterinaire  Pub- 
lique,  1891,  No.  4). 


—  8o  — 

excited  by  tuberculin  when  freed  from  glycerin.  This 
explanation  is  at  the  best  a  strange  one. 

How  can  the  positive  chimiotactic  power  of 
tuberculin,  i.e.,  the  action  which  it  may  exercise  in 
certain  conditions  on  the  leucocytes,  explain  the  exu- 
dative inflammation  produced  by  tuberculin,  not  at 
the  spot  of  inoculation,  but  at  the  place  of  election  ? 
This  impossible  comparison  can  only  be  understood 
by  calling  to  mind  the  confusion  which  exists  in  cer- 
tain microscopists  with  regard  to  the  difference  be- 
tween inflammation  and  the  emigration  of  leucocytes, 
the  latter  being  only  an  element  more  or  less  constant 
of  inflammation,  and  indeed  oftener  found  apart 
from  it.* 

The  explanation  of  the  elective  action  of  tuber- 
culin is  not,  however,  so  difficult,  and  it  has  already 
been  sketched  by  Koch.  The  acute  inflammation  is 
in  general  the  consequence  of  a  lesion  or  rapid  de- 
struction of  the  cells  and  tissues  of  the  animal.  When 
injected  in  sufficient  concentration,  the  modified 
poisons  induce  cellular  lesions  and  exudative  inflam- 
mation at  the  place  of  their  injection.  Appropriately 
diluted,  they  are  inoffensive  to  the  normal  cells  of 
the  economy,  and  affect  only  the  tissues  already  dis- 
eased, around  which  is  then  produced  the  inflamma- 
tory process.  This  explanation  agrees  well  with  the 


*This  confusion  has  even  been  made  the  basis  of  a  new 
theory  of  inflammation. 


—  8i  — 

property  of  other  substances  besides  tuberculin  to 
provoke  reaction  in  the  tuberculous,  and  with  the 
property  of  tuberculin  of  provoking  also  reaction 
around  certain  lesions  not  tuberculous. 

The  relations  between  the  microbian  poisons  and 
the  animal  economy  have  been  chiefly  studied  from 
the  point  of  view  of  immunity.  When  vaccination  by 
means  of  the  soluble  products  of  bacteria  was  first 
demonstrated,  the  very  natural  idea  arose  that  this 
immunity  was  acquired  by  the  habituation  of  the  ani- 
mals to  the  poison.  According  to  the  different  hypoth- 
eses respecting  immunity,  this  habituation  was  sup- 
posed to  affect  either  the  entire  animal  economy,  or 
only  certain  of  its  cells.  This  idea  was,  moreover,  con- 
firmed by  the  facts  connected  with  one  of  the  first 
chemical  vaccinations  known.  Beumer  noted  that  he 
could  easily  habituate  mice  to  progressively  increas- 
ing doses  of  the  sterilized  cultures  of  the  typhoid  bacil- 
lus. These  mice,  having  become  accustomed  in  this 
way  to  the  poison,  were  able  to  sustain  without  injury 
the  inoculation  of  large  quantities  of  the  living  bacilli 
— a  procedure  which  proved  speedily  fatal  to  mice  not 
vaccinated.*  Similar  facts  have  been  noted  by  Foa 
and  Bonome  in  the  chemical  vaccination  of  hares 
against  the  proteus  vulgaris.f 

*  Beumer:  Der  derzeitige  Shandpunkt  der  Schutzim- 
pfungen,  pp.  4-6.  Wiesbaden,  1887. 

f  Foa  and  Bonome:  Ueber  Schutzimpfungen  (Zeitschrift 

f.  Hygiene,  t.  v,  p.  415). 
6  ooo 


—    82    — 

But  in  my  own  researches  on  vaccination  against 
the  avicide  vibrio  found  by  me,  I  have  met  with 
facts  which  overthrow  this  theory  of  habituation. 
The  cultures  of  the  avicide  vibrio,  sterilized  at 
120°  C.  and  very  toxic  for  guinea-pigs,  easily  vac- 
cinate them  against  infection  by  the  living  vibrio. 
But  after  being  once  vaccinated  the  guinea-pigs 
do  not  become  more  resistant  to  the  toxic  action 
of  the  vaccine — /.  <?.,  to  massive  doses — than  the  non- 
vaccinated.  The  fever  which  appeared  after  the 
injection  of  small  doses  was  in  them  identical  with 
that  of  the  control-animals.  The  hypothermia  fol- 
lowing larger  doses  had  the  same  characters.  The 
exudative  inflammation  which  the  vaccine  provokes 
at  the  point  of  inoculation  was  not  modified  in  conse- 
quence of  the  vaccination  of  the  guinea  pigs.  The 
phenomena  of  positive  or  negative  chimiotaxy  de- 
pended on  the  dose  of  vaccine  introduced,  but  not  on 
the  immunity  or  non-immunity  of  the  guinea-pigs. 
In  short,  the  behavior  of  the  guinea-pigs  toward  the 
poison  of  the  chemical  vaccine  was  in  no  way 
changed  by  the  vaccination.  Moreover,  we  have 
found  that  the  volatile  products  of  the  non-toxic 
cultures  can  also  vaccinate.  We  have  found  this 
same  failure  to  undergo  habituation  in  animals 
subjected  to  cholera  vaccine,*  to  pyocyanic  vac- 

*The  facts  which  we  have  found  have  been  confirmed 
by  many  observers.  See  especially  Hernandez,  "Chemical 
Vaccinations,"  in  C.  R.  de  la  Soc.  de  Biologic,  July  4,  1891; 


—  83  - 

cination  (Charrin  and  I),  to  hog-cholera  vaccine 
(Selander).f 

All  these  facts  show  that  the  chemical  vaccina- 
tion of  animals  does  not  depend  on  their  becoming 
accustomed  to  the  vaccine.  They  may  be  explained 
by  the  hypothesis  that  in  our  vaccine  are  mingled  the 
poison,  and  the  vaccinant  substance  distinct  from  the 
poison.  This  hypothesis  of  the  distinction  between 
the  toxic  substance  and  the  vaccinant  substance, 
formulated  first  by  Bouchard, J  may  account  for  the 
mechanism  of  vaccination,  but  it  is  manifestly  insuffi- 
cient, for  it  does  not  explain  the  positive  cases  of  re- 
sistance to  the  poisons  by  the  vaccinated  animals  in 
the  experiments  of  Beumer,  Foa,  and  Bonome,  and 
others  of  whom  we  shall  speak  presently. 

The  complete  explanation  of  this  difficulty  can 
only  be  obtained  by  the  aid  of  the  notion  of  the 
plurality  of  the  microbian  poisons.  We  have  estab- 
lished the  distinction  between  the  natural  or  primary 
poisons  of  the  microbes,  and  their  artificial  or  modi- 


and  Metschnikoff  and  Roudenko,  "  Researches  on  Habitua- 
tion  to  the  Microbian  Products  "  (Ann.  de  1'Instit.  Pasteur, 
1891,  No.  9). 

f  Gamaleia:  On  Cholera  Vaccination  (C.  R.  de  la  Soc. 
de  Biologic,  Nov.  30,  1891).  Charrin  and  Gamaleia:  Vacci- 
nation et  Accoutumance  (C.  R.  de  la  Soc.  de  Biologic,  July 
5,  1890).  Selander:  Study  of  Hog-Cholera  (Ann.  de  Tlnst. 
Pasteur,  1890,  p.  545). 

t  Bouchard:  Therapeutique  des  Malades  Infectieuses, 
p.  137.  Paris,  1889. 


_  84- 

fied  poisons.*  With  this  distinction,  all  the  foregoing 
contradictions  are  cleared  up.  If  our  animals  vac- 
cinated against  the  infection  did  not  become  refrac- 
tory to  the  poisons  contained  in  the  chemical  vac- 
cines, this  was  owing  to  the  fact  that  the  latter 
poisons  are  artificial  products  and  have  nothing  to  do 
with  the  infection  and  the  natural  poisons  of  the  bac- 
teria. 

In  fact,  towards  these  natural  poisons  the  vac- 
cinated animals  become  quite  as  refractory  as  towards* 
the  microbes  themselves.  This  last  fact  was  first 
established  by  the  researches  of  Behring  and  Kitasata 
in  reference  to  tetanus  and  diphtheria,  f  G.  and  F. 
KlempererJ  have  noticed  the  same  resistance  to  the 
pneumonic  poison  on  the  part  of  animals  vaccinated 
against  the  streptococcus  lanceolatus  of  pneumonia.  In 
cholera,  lastly,  we  have  seen  that  vaccinated  dogs 
become  refractory  to  the  natural  poison  of  cholera 
and  not  to  the  modified  poison.  Quite  recently,  also, 


*GamaleIa:  On  the  Diarrhoeic  Action  of  the  Cultures  of 
Cholera  (C.  R.  de  1'Acad.  des  Sciences,  March  24,  1890). 
Gamalela:  On  the  Two  Poisons  of  Cholera  (Bulletin  Medical,. 
April,  1890). 

f  Behring  and  Kitasato:  Facts  as  to  Immunity  from, 
Tetanus  and  Diphtheria  (Deutsche  Medic.  Wochensch.,  1890^ 
No.  49)- 

$G.  and  F.  Klemperer:  Immunity  against  Pneumo- 
coccus  Infection  (Berlin.  Klin.  Wochensch.,  1891,  Nos.  34 
and  35). 


-  85  — 

in  respect  to  typhoid  fever,  the  researches  of  Beumer 
and  Peiper  on  the  resistance  of  vaccinated  animals  to 
the  poison  have  been  confirmed  by  those  of  Brieger, 
Kitasato,  and  Wassermann.* 

We  may  consequently  regard  it  as  a  very  general 
fact  that  animals  refractory  to  infection  by  a  microbe 
also  resist  intoxication  by  the  natural  poison  of  this 
microbe.  This  principle  finds  verification,  not  only 
in  reference  to  artificial  immunity  conferred  by  vac- 
cination, but  also  to  natural  immunity.  Thus,  for 
instance,  rats  which  are  refractory  to  the  diphtheritic 
infection,  support  without  difficulty,  according  to 
Roux  and  Yersin,  the  injection  of  enormous  doses  of 
the  poison  of  diphtheria. 

We  may  conclude  that  immunity  is  linked  to  the 
resistance  of  animals  to  the  microbian  poisons. 

To  what  is  this  resistance  due?  We  have  already 
•seen  that  it  is  not  the  result  of  accustomance,  for  by 
a  chemical  vaccine  which  does  not  contain  any  natural 
poison  we  vaccinate  against  a  natural  poison. 

Behring  and  Kitasato  have  shown  that  in  respect 
to  tetanus  and  diphtheria,  acquired  immunity  and  re- 
sistance to  the  poison  are  linked  to  the  power  of  the 
serum  of  the  vaccinated  animals  to  destroy  even  in 
vitro  the  corresponding  poison.  This  remarkable  dis- 
covery has  been  since  then  often  confirmed  in  its  ap- 


*  Brieger,    Kitasato,    and    Wassermann,    Zeitschrift    f. 
Hygiene,  t.  xii,  p.  137. 


—  86  — 

plication  to  diphtheria  and  tetanus,  and  also  to  other 
poisons.* 

At  the  same  time,  this  antitoxic  property  of  the 
serum  cannot  be  the  cause  of  the  resistance  of  ani- 
mals to  the  microbian  poisons;  it  is  rather  the  con- 
sequence, for  animals  naturally  refractory  do  not 
ordinarily  possess  this  antitoxic  property  of  the 
serum — they  do  not  acquire  it  until  after  having  de- 
stroyed the  poisons  in  their  bodies. f  The  immunity 
cannot  therefore  depend  on  the  antitoxic  power  of 
the  serum;  it  can  only  be  characterized  by  the  pos- 
sibility of  acquiring  this  antitoxic  power  after  the  in- 
troduction of  the  poison. 

But  the  question  as  to  the  nature  of  immunity  is 
foreign  to  our  book,  and  we  cannot  enter  into  expla- 
nations respecting  its  physiological  mechanism. 

To  sum  up,  briefly,  all  that  we  know  as  to  the  gen- 
eral toxicology  of  the  microbes,  we  shall  see  that  our 
hypothesis  as  to  the  chemical  nature  of  the  bacterial 
poisons,  in  absence  of  a  direct  demonstration  which 
can  only  be  made  with  the  poisons  isolated  in  a 
state  of  purity,  is  found  confirmed  by  many  indirect 
proofs. 

That  the  microbian  poisons  belong  to  the  class"  of 
nucleo-albumins,  is  in  agreement  with  their  extreme 


*Ehrlich:  Experimentelle  Untersuchungen  tiber  Immu- 
nitat  (Deutsche  Medic.  Woch.,  1891,  Nos.  32  and  44). 

fGamaleia:  L'immunisation  (Gaz.  Hebdom.  de  Med. 
et  de  Chir.,  Nov.  21,  1891). 


—  87  — 

fragility,  and  the  facility  with  which  they  give  rise  to 
the  modified  poisons  on  the  one  hand,  to  the  pto- 
maines on  the  other.  This  idea  is  also  confirmed  by 
what  we  know  about  the  source  of  the  poisons  of  the 
microbian  bodies,  which  are  of  nuclear  origin.  It  is, 
lastly,  compatible  with  the  plurality  of  poisons  asso- 
ciated together  and  produced  by  the  same  bacteria — a 
plurality  postulated  by  the  physiological  study  of 
these  poisons. 

We  shall  speak  only  of  the  principal  facts  which 
confirm  our  hypothesis. 


Third  Part:  The  Special  Toxicology  of 
the  Microbes. 


CHAPTER  IX. 

THE    POISONS   OF   TETANUS. 


SUMMARY. — Researches  of  Brieger,  Kitasato  and  Weyl, 
Knud  Faber,  Tizzoni  and  Cattani,  Brieger  and 
Frdnkel,  Vaillard  and  Vincent  and  Kitasato,  on 
the  Properties  of  the  Tetanus  Poison —  The  Investi- 
gations of  Bruschetini  and  of  Camara  Pestana  on 
its  Diffusion  in  the  Animal  Body — Vaccination 
against  Tetanus:  Behring  and  Kitasato —  The  New 
Notions  Contributed  by  these  Authors. 

We  begin  our  exposition  by  the  study  of  the 
poisons  produced  by  the  bacillus  of  tetanus.  These 
poisons  are  not  the  first  known,  but  they  are  in  many 
respects  the  best  studied,  of  the  microbian  poisons. 

Brieger  was  the  first  to  investigate  the  toxic 
products  formed  by  the  bacillus  of  tetanus.  In  the 
cultures  of  this  bacillus,  as  well  as  in  the  amputated 
arm  of  a  patient,  he  found  several  characteristic 
ptomaines.  There  was  tetanm,  which  has  the  prop- 
erty of  provoking  in  mice,  even  in  very  small  doses, 


_  89  - 

attacks  of  trismus  and  of  tetanus  ending  in  death; 
tetanotoxin,  which  provokes  convulsive  paroxyms, 
followed  by  complete  paralysis;  spasmotoxin,  which 
causes  cramps;  and  lastly,  toxin,  which  has  the  prop- 
erty of  stimulating  the  salivary  and  lachrymal  secre- 
tion. 

The  researches  of  Brieger  date  from  a  period 
when  we  did  not  know  how  to  cultivate  the  tetanic 
bacillus  in  a  state  of  purity.  Since  then,  Kitasato  has 
succeeded  in  making  pure  cultures  of  tetanus.  One 
may  ask  if  in  the  pure  cultures  the  ptomaines  of 
Brieger  may  be  found  ?  Kitasato  and  Weyl  have 
put  to  themselves  this  question.  They  have  suc- 
ceeded, by  employing  Brieger's  method,  in  finding  in 
notable  quantity  tetanin  and  traces  of  tetanotoxin. 
Among  the  volatile  products  of  tetanus  cultures  they 
have  found  H2S,  butyric  acid,  indol,  and  phenol. 
But  the  two  bases  found  were  little  toxic,  and  quite 
insufficient  to  explain  the  disease.  Tetanin  does 
not  provoke  convulsions  and  salivation  in  mice  except 
when  given  in  very  considerable  doses,  and  spasmo- 
toxin causes  only  paralysis.* 

It  was  evident  (and  this  is  the  conclusion  of 
these  authors)  that  the  true  tetanus-poison  must  be 
found  elsewhere. 


*  Brieger:  Ueber  Ptomaine,  iii,  Theil.  Berlin,  1886. 
Kitasato  and  Weyl:  Zur  Kentniss  der  Anaeroben  (Zeitschrift 
f.  Hygiene,  t.  viii,  p.  404). 


—  90  — 

Moreover,  already  in  1890  Faber  had  found  in 
tetanus  cultures  a  poison  of  quite  another  nature  than 
the  ptomaines  of  Brieger.  In  filtering  cultures  of  the 
tetanus  bacillus  through  the  Chamberland  filter, 
Faber  obtained  liquids  deprived  of  bacilli,  but  repro- 
ducing in  animals  all  the  symptoms  of  tetanus  in  the 
same  degree  as  the  tetanus  bacilli  themselves.  In- 
toxication by  the  poison  resembled  infection  by  the 
bacillus  itself  in  the  two  following  respects:  (i)  both 
demand  a  certain  time  for  their  manifestation — a 
period  of  incubation  for  the  intoxication  as  for  the 
infection;  (2)  when  introduced  into  the  stomach,  the 
chemical  poison  is  quite  as  inoffensive  as  the  living 
bacillus.  Faber  also  observed  that  the  toxicity  of  the 
filtered  cultures  was  completely  destroyed  by  a  five- 
minutes  heating  to  65°  C.,  and  even  by  the  addition  of 
alcohol. 

He  concludes  that  the  tetanus  poison  is  not  a 
ptomaine,  but  rather  a  toxic  diastase  like  that  of 
jequirity  or  that  of  diphtheria.  Faber  had  not  at  his 
disposal  pure  cultures  of  tetanus.* 

But  these  results  were  soon  confirmed  by  Tizzoni 
and  Cattani.  These  authors  have  found  that  the 
tetanus  bacillus  produces  its  poison  in  pure  cultures 
on  gelatin,  and  not  in  such  as  are  made  in  bouillon. 
The  toxic  substance  of  the  filtered  cultures  is  modi- 


*  Knud  Faber:    On  the  Pathogeny  of  Tetanus  (Berlin. 
Klin.  Woch.,  1890,  No.  31). 


fied  by  precipitation  by  alcohol.  It  does  not  dialyze, 
and  this  has  enabled  the  authors  to  isolate  it  in  the 
following  way:  Filtered  cultures  are  supersaturated 
by  ammonium  sulphate  in  excess;  a  precipitate  is 
formed,  which  contains  the  toxic  substance;  this 
precipitate  is  dissolved  in  water,  freed  from  salts  by 
dialysis,  and  evaporated  in  a  vacuum  at  a  low  tem- 
perature. The  toxic  substance  obtained  in  this  way 
presents  itself  as  a  yellow  body,  of  crystalline  aspect. 
It  is  rendered  completely  inactive  by  thirty  minutes' 
heating  up  to  60°  C.  One  hour's  heating  at  55°  C. 
reduces  its  toxicity.  The  alkalies,  carbonic  acid, 
organic  acids,  and  dilute  mineral  acids,  do  not  modify 
it.  The  concentrated  mineral  acids  destroy  its  toxi- 
city. 

As  to  the  nature  of  this  tetanus  poison,  Tizzoni 
and  Cattani  note  numerous  similarities  with  the 
soluble  ferments.  In  gelatin  the  tetanus  bacillus 
secretes  a  peptic  ferment  which  it  does  not  produce 
in  bouillon,  and  all  the  influences  which  modify  the 
tetanus  poison  destroy  also  the  peptic  diastase.* 

In  their  researches  on  the  bacterial  poisons, 
Brieger  and  Frankel  speak  of  the  toxalbumin  of 
tetanus.  This  toxic  substance  was  soluble  in  water. 
Its  parent  cultures  were  developed  in  sweetened 


*  Tizzoni  and  Cattani:  Sul  Veleno  di  Tetano  (Riforma 
Medica,  1890,  No.  128).  Tizzoni  and  Cattani:  Experimentelle 
Untersuchungen  liber  das  Tetanusgift  (Archiv  f.  Exper. 
Pathol.,  Bd.  xxvii). 


bouillon,  then  filtered  through  the  Chamberland  filter, 
reduced  by  evaporation  in  a  vacuum,  and  precipitated 
by  absolute  alcohol.*  Vaillard  and  Vincent  have  ob- 
tained in  tetanus  cultures,  made  in  bouillon  and  fil- 
tered, an  extremely  active  poison,  of  which  -fa  and  y^ 
c.c.  sufficed  to  killed  a  guinea-pig.  They  have  con- 
firmed most  of  the  foregoing  data:  thus,  for  instance, 
the  feeble  resistance  of  the  poison  to  heat,  and  its  inac- 
tivity in  the  digestive  tube.  Among  the  new  data  of 
their  work,  I  may  cite  especially  the  following  facts: 
Direct  solar  light  destroys  rapidly,  in  free  air,  the 
toxicity  of  the  filtered  liquid.  Acidification  does 
not  modify  its  toxic  power.  Absolute  alcohol  does 
not  alter  the  toxic  substance;  it  precipitates  it  in 
part,  and  the  product  of  this  precipitation  is  tetani- 
gen.  The  tetanus  poison  adheres  to  certain  precipi- 
tates which  are  produced  in  the  liquids  in  which  it  is 
contained:  thus  it  is  that  the  precipitates  of  phos- 
phate of  lime  or  of  alum  carry  down  a  part  of  this 
active  substance,  but  this  precipitation  is  quite  in- 
complete. Vaillard  and  Vincent  believe  that  the 
tetanus  poison  belongs  to  the  diastases,  f 

Kitasato  has  made  a  detailed  study  of  the  tetanus 
poison.  He  has  determined  the  exact  length  of  time 

*Briegerand  Frankel:  Untersuchungen  ttber  Bacteri- 
engifte  (Bui.  Kl.  Woch.,  1890,  Nos.  n  and  12). 

f  Vaillard  et  Vincent:  Le  Poison  Tetanique  (C.  R.  de  la 
Societe  de  Biologic,  1890,  Nov.  13).  "Vaillard  et  Vincent: 
Contribution  a  1'Etude  du  Tetanos  (Annales  de  1'Institut 
Pasteur,  1891,  No.  i). 


—  93  ~ 

necessary  for  the  different  physical  influences  (light, 
heat,  etc.),  or  such  chemical  influences  as  acids  and 
alkalies  to  destroy  the  tetanus  poison.  He  has  found 
that  absolute  alcohol  added  in  sufficient  quantity  not 
to  leave  the  poison  in  solution  destroys  it.  In  general, 
Kitasato  has  not  found  the  means  of  isolating  the 
tetanus  poison  from  the  liquid  in  which  it  is  con- 
tained.* 

Here  end  our  sources  of  knowledge  as  to  the 
chemical  nature  of  the  tetanus  poison.  As  to  the 
mode  of  its  action  on  the  animal  organism,  we  have 
not  made  much  advancement  since  the  researches 
of  Knud-Faber.  We  know  that  the  poison  repro- 
duces exactly  all  the  symptoms  by  tetanus  in  the 
same  way  as  we  obtain  them  with  the  bacillus  itself. 
These  phenomena  of  systemic  poisoning  do  not  ap- 
pear immediately  after  the  injection  of  the  toxic 
liquid,  but  after  a  certain  time  of  incubation,  which 
diminishes  as  the  dose  or  the  toxicity  of  the  in- 
jected filtered  liquid  augments. 

The  morbid  symptoms  begin,  in  the  case  of  sub- 
cutaneous injection  of  the  liquid,  at  or  near  the  point 
of  inoculation.  The  first  contractions  appear  in  the 
muscles  nearest  the  seat  of  inoculation.  Bruschetini 
has  studied  the  diffusion  of  the  tetanus  poison  in  the 
animal  organism;  he  finds  that  it  takes  place  chiefly 


*  Kitasato:    Experimentelle    Untersuchungen  tiber  das- 
Tetanusgift,  t.  x,  p.  267. 


—  94  — 

along  the  nervous  system.  He  made  use  of  filtered 
cultures  of  tetanus.* 

Camara  Pestana,  working  with  filtered  cultures, 
has  carried  out  the  following  conclusions: 

i st.  The  absorption  of  the  toxine  takes  place  by 
the  blood. 

2d.  The  lungs,  the  spleen,  the  kidneys,  but  prin- 
cipally the  liver,  take  from  the  blood  the  toxic  princi- 
ple, and  retain  it. 

3d.  The  toxine  is  not  appreciably  eliminated  by 
the  urine. 

4th.  Despite  the  predominence  of  the  neuro- 
muscular  phenomena,  no  one  has  yet  succeeded  in 
detecting  the  presence  of  the  toxine  in  the  nervous 
and  muscular  tissue,  and  all  the  experiments  made 
with  these  tissues  have  given  negative  results,  f 

The  presence  of  the  toxine  in  the  blood  of  per- 
sons affected  with  tetanus  has  also  been  noted  by 
Bruschetini,  Kitasato,  and  Nissen. 

The  physiological  study  of  the  influence  of  the 
tetanus  poison  on  the  different  parts  of  the  neuro- 
muscular  apparatus  has  not  yet  been  made,  and  we  do 
not  know  how  this  poison  acts.  Faber  has  presented 


*  Bruschetini:  Recherches  Preliminaires  sur  la  Diffu- 
sion du  Poison  du  Tetanus  dans  1'  Organisme  (Annales  de 
Micrographie,  1890,  Nov.  20). 

f  Camara  Pestana:  Diffusion  du  Poison  du  Tetanos  dans 
1'Organisme  (C.  R.  de  la  Societe  de  Biologic,  1891,  June  27). 


—  95  — 

the  hypothesis  that  it  affects  the  terminal  nerve-plates, 
like  curare. 

With  regard  to  the  subject  of  immunity  from 
tetanus,  there  are  some  important  details.  At  first, 
all  the  experimenters  arrived  at  constantly  negative 
results  in  trying  to  vaccinate  animals  against  tetanus. 
At  length,  Behring  and  Kitasato  announced  that  they 
had  succeeded  in  rendering  animals  immune  by  vac- 
cination. The  animals  vaccinated  against  tetanus  ex- 
emplified by  their  behavior  the  following  facts,  which 
were  quite  new  and  important: 

1.  The  hares  which  were  vaccinated  and  resisted 
the  inoculation  of  the  living  tetanus  bacillus,  were 
also  refractory  to  the  injection  of  the  chemical  poison 
produced  by  this  bacillus. 

2.  This  insensibility  of  the  vaccinated  animals  to 
the  poison  is  not  due  to  an  habituation  of  their  organ- 
ism to  the  poison,   but   to    its    destruction   in   their 
bodies.    In  fact,  the  blood,  and  more  particularly  the 
serum  of  the   blood,  of  the  vaccinated  hares,  when 
mixed  with  the  tetanus  poison,  even  in  small  propor- 
tion, destroys  it.     The  blood  of  animals  not  vaccin- 
ated does  not  possess  this  antitoxic  property. 

3.  The   serum  of  vaccinated   animals  possesses 
not  only  in  vitro  the  antitoxic  property,  it  also  exer- 
cises it  in  the  bodies  of  other  animals.     For  the  serum 
of  vaccinated  hares,  when  injected  in  mice,  renders 
them  refractory  to  subsequent  inoculation  of  the  bacil- 
lus or  of  the  tetanus-poison.     It  may  even  cure  tetanus 
in  mice  already  sick. 


_  96  - 

We  see  that  this  work  of  Behring  and  Kitasato 
contains  three  new  and  important  notions:  Vaccina- 
tion against  tetanus;  the  antitoxic  action  of  the  animal 
humors;  and  immunization  by  these  humors.  These 
three  notions  have  been  controlled  and  studied  since 
then  by  many  experimenters,  and  in  general  have 
been  found  perfectly  exact.  In  the  following  chap- 
ter we  shall  state  the  results  acquired  by  these  stud- 
ies.* 


*  Behring  et  Kitasato:  Ueber  das  Zustande-Kommen 
der  Diphteric  immunitat  und  der  Tetanus  immunitat  bei 
Thieren  (Deutsche  Medic.  Wochenschrift,  1890,  No.  49). 


CHAPTER  X. 

THE    POISONS  OF  TETANUS— (Continued). 


SUMMARY.  —  Vaccination  against  Tetanus  is  Obtained 
by  Tizzoni,  Cattani,  and  Vaillard — The  Researches 
of  Kitasato;  of  Behring;  of  Brieger,  Kitasato, 
and  Wassermann — Immunization — The  Labors  of 
Tizzoni,  Cattani,  and  Vaillard —  The  Researches  of 
Ehrlich;  of  Brieger  and  Ehrlich;  of  Brieger  and 
Frank — Application  of  the  Method  of  Behring  and 
Kitasato  to  the  Treatment  of  Tetanus  in  Man. 

We  must  remark  that  following  the  epoch  of  his 
publication  with  Behring,  Kitasato  did  not  possess  a 
method  of  vaccinating  animals  against  tetanus.  He 
had  only  at  his  command  certain  animals  which  were 
shown  to  be  fortuitously  vaccinated;  for  a  long  time 
he  had  been  able  to  give  only  erroneous  procedures 
for  this  vaccination. 

But  independently  of  these  researches  other  ex- 
perimenters have  succeeded  in  vaccinating  animals 
against  tetanus. 

Tizzoni  and  Cattani  have  vaccinated  pigeons  and 
dogs  against  tetanus  by  injecting  little  doses  of  the 
living  cultures  of  tetanus.  On  these  vaccinated  ani- 
mals they  have  been  able  to  confirm  the  fundamental 

7  OOO 


propositions  of  Behring  and  Kitasato  respecting  anti- 
toxic reactions  and  immunization.*  Vaillard,  in  util- 
izing the  method  of  C.  Frankel  (which  we  shall  de- 
scribe under  the  head  of  Diphtheria),  has  been  able 
to  vaccinate  hares.  The  method  of  Vaillard  consists 
in  heating  for  an  hour,  at  60°  C.,  filtered  cultures  of 
tetanus.  The  tetanus  poison  is  modified  by  heat  to 
the  point  of  being  unable  longer  to  provoke  tetanus 
even  when  injected  in  large  doses.  But  it  acquires,  by 
heating,  the  property  of  conferring  immunity.  If  we 
heat  the  filtered  cultures  to  65°  C.,  they  no  longer 
confer  immunity.! 

KitasatoJ  described  later  a  process  for  vaccin- 
ating against  tetanus.  It  consists  in  inoculating  hares 
with  the  living  and  virulent  cultures  of  tetanus,  mixed 
with  progressively  decreasing  doses  of  bichloride  of 
iodine.  But  the  results  were  first  but  little  favorable, 
for  out  of  fifteen  hares  he  succeeded  in  vaccinating 
only  six.  Since  then,  however,  the  process  with 
ter-chloride  of  iodine  (IC13)  has  been  sensibly  im- 
proved at  Koch's  Institute  by  Behring,§  so  far  as  to 


*Tizzoni  and  Cattani:  Ueber  die  Art  einem  Thiere  die 
Immunitat  gegen  Tetanus  zu  tibertragen  (Centralblatt  f.  Bac- 
teriologie,  t.  ix,  No.  6). 

f  Vaillard:  Immunite  centre  le  Tetanos  (C.  R.  de  la 
Soc.  de  Biol.,  1891,  Feb.  21). 

\  Kitasato:  Experimentelle  Untersuchungen  liber  das 
Tetanusgift  (Zeitschrift  f.  Hygiene,  t.  x,  p.  267). 

§  Behring:  Ueber  Immunisirung  und  Heilung  von  Ver- 
auchtieren  beim  Tetanus  (Zeitschrift  f.  Hygiene,  t.  xii,  p.  45). 


—  99  — 

enable  us  to  vaccinate  without  great  danger  all  the 
animals  under  experimentation:  mice,  hares,  sheep, 
and  horses.  Still  this  process  makes  the  animals 
quite  sick  for  some  time.  Lastly,  Brieger,  Kitasato, 
and  Wassermann  have  found  at  Koch's  Institute  a 
method  of  vaccination  perfectly  inoffensive.  Resum- 
ing the  already  antiquated  researches  of  Wooldridge 
on  cultures  of  the  microbes  in  extract  of  thymus,  and 
following  exactly  his  indications,  these  authors  have 
made,  with  the  thymus,*  culture-media  which  have 
given  them  interesting  results  with  the  different 
microbes,  especially  with  the  bacillus  of  tetanus. 
These  tetanus  cultures  made  in  extract  of  thymus  are 
asporogenous  and  very  slightly  toxic.  On  the  other 
hand,  the  extract  of  thymus  mixed  with  a  filtered  cul- 
ture of  tetanus  made  in  ordinary  bouillon,  and  very 
toxic,  destroys  little  by  little  the  toxicity  of  the 
culture  product.  In  employing  these  mixtures  when 
two  days  old  in  progressively  increasing  doses,  the 
authors  have  succeeded  in  vaccinating  without  dan- 
ger and  without  apparent  trouble,  animals,  such  as 
mice,  the  most  sensitive  to  tetanus.f 

Vaillard  has  recently  come  back  to  the  question 


*  Wooldridge:  Versuche  iiber  Schuzimpfung  auf  Chem- 
ischen  Wege  (Archiv  f.  Anatomic  und  Physiologic,  Physio- 
logische  Abtheilung,  1888,  p.  527). 

f  Brieger,  Kitasato,  and  Wassermann:  Ueber  Immuni- 
tat  und  Giftfestigung  (Zeitschrift  f.  Hygiene,  t.  xii,  p. 
1887). 


100    

of  vaccination  against  tetanus.*  Besides  his  old 
method  of  vaccination  by  cultures  filtered  and  heated 
to  60°  C.,  he  indicates  two  new  processes:  one  of  these 
is  very  nearly  like  that  of  Behring  and  Kitasato,  and 
consists  in  injecting  in  animals  tetanus-cultures  to 
which  iodine  solution  has  been  added;  the  other  is- 
identical  with  that  of  Tizzoni  and  Cattani — inocula- 
tion with  extremely  small  doses  of  the  virus.  Behring 
has  also  utilized  with  success  this  process  of  vaccina- 
tion after  dilution. 

It  will  be  seen  that  this  question  of  vaccination 
has  gone  through  three  successive  stages.  At  first, 
experimenters  only  succeeded  in  vaccinating  a  certain 
proportion  of  the  animals  under  trial,  while  the  others 
died  in  consequence  of  the  manipulations.  Then  they 
obtained  more  constant  results,  but  the  vaccinated 
animals  suffered  more  or  less  severely  in  their  general 
health  (local  tetanus,  fever,  and  emaciation).  At  last 
they  found  a  method  both  sure  and  inoffensive  (Brie- 
ger,  Kitasato,  and  Wassermann).  The  antitoxic  prop- 
erties of  the  serum  of  vaccinated  animals  were  soon 
found  by  all  the  experimenters. 

Tizzoni  and  Cattani  have  made  a  special  study 
of  the  antitoxic  substance  of  serum.  They  have 
found  that  their  antitoxine  is  enfeebled  by  heating  to 
65°  C.  half  an  hour,  and  completely  destroyed  by 
heating  it  the  same  length  of  time  at  68°  C.  (the 


*  Vaillard,  Annales  de  1'Institut  Pasteur,  1892,  No.  4. 


101    

temperature  of  coagulation  of  the  serum).  They 
have  found  also  that  this  antitoxic  substance  does 
not  dialyze;  that  it  is  destroyed  very  quickly  by  hy- 
drochloric acid,  by  lactic  acid  in  great  excess,  and 
by  the  alkalies.  It  is  precipitated  by  ammonium  sul- 
phate in  supersaturation,  and  also  by  absolute  alco- 
hol. From  this  latter  precipitate  it  can  be  extracted 
by  water  or  by  glycerin.* 

Vaillard  has  studied  especially  the  relations  be- 
tween the  immunity  of  animals  and  the  antitoxic 
property  of  this  serum.  He  has  been  led  by  his  re- 
searches to  deny  the  possibility  of  explaining  immu- 
nity by  the  antitoxic  property  of  serum.  In  animals 
naturally  refractory,  as,  for  instance,  hens,  the  serum 
in  the  natural  state  is  not  antitoxic;  it  only  acquires 
this  property  after  the  injection  of  a  large  dose 
of  tetanus  poison.  Likewise,  in  the  vaccinated  ani- 
mals, one  may  not  find  the  antitoxic  state  of  the 
serum.  This  state  only  appears  as  the  result  of  the 
action  of  large  doses  of  the  soluble  products  of  the 
tetanus  bacillus.  The  antitoxic  action  of  the  serum  is 
only  a  contingent  property  of  the  refractory  animals. 
It  cannot,  therefore,  serve  to  explain  natural  or  ac- 
quired immunity.  We  must  add  that  Vaillard  did 
not  succeed  in  curing  tetanus  by  the  serum  of  his  re- 
fractory animals.  He  found  also  that  the  spleen  and 


*Tizzoni  et  Cattani:  Ueber  die  Eigenschaften  des  Te- 
tanus Antitoxins  (Centr.  f.  Bacteriologie,  t.  ix,  No.  21). 


IO2 


aqueous  humor  of  the  vaccinated  animals  did  not 
possess  antitoxic  properties.  Their  muscles,  however, 
possess  these  properties,  at  least  in  vitro* 

Apropos  of  the  spleen,  we  must  speak  of  the  re- 
searches of  Tizzoni  and  Cattani,  who  have  found  that 
it  is  impossible  to  vaccinate  against  tetanus  animals 
that  are  deprived  of  their  spleen. 

The  third  thesis  of  Behring  and  Kitasato — vac- 
cination and  cure  by  the  serum  of  vaccinated  animals 
— has  also  been  contested.  While  succeeding  in  pre- 
venting the  disease  by  the  serum,  neither  Tizzoni  and 
Cattani,  nor  Vaillard,  succeeded  in  curing  it. 

Many  other  contradictions  have  been  affirmed 
against  the  general  theory  of  immunization — contra- 
dictions due  for  the  most  part  to  the  partisan  influ- 
ence of  the  old  theories  of  immunization;  but  all 
these  contradictions  have  received  their  quietus, 
thanks  especially  to  the  interesting  researches  of 
Ehrlich.f  Experimenting  with  abrine  and  ricine,  two 
toxic  albuminoids  of  vegetable  origin,  Ehrlich  dem- 
onstrated numerically  that  the  immunity  of  animals 
against  the  poison  is  not  an  invariable  quantity,  but 
may  have  very  different  degrees.  He  demonstrated, 
moreover,  that  with  the  degree  of  the  immunity 


*  Vaillard:  Property  of  the  Serum  of  Animals  Refrac- 
tory to  Tetanus  (C.  R.  de  la  Societe  de  Biologic,  June  6, 
1891). 

f  Ehrlich,  Deutsche  Med.  Wochenschrift,  1891,  Nos.  32 
and  44. 


—  103  ~ 

acquired  by  the  vaccinated  animals  varies  also  the 
antitoxic  and  immunizing  power  of  their  serum. 

Thus  were  cleared  up  all  the  previous  difficulties; 
the  experimenters  who  could  not  cure,  but  who  vac- 
cinated only  with  the  serum  of  vaccinated  animals,  had 
evidently  not  obtained  in  those  animals  the  grade  of 
immunity  sufficient  for  their  serum  to  be  curative.  In 
fact,  in  continuing  their  experiments,  Tizzoni  and 
Cattani,  and  Vaillard,  have  perfectly  succeeded  in 
curing  their  tetanic  animals  with  the  serum  of  the 
vaccinated  animals.  At  the  present  time,  there  re- 
mains no  longer  any  doubt  as  to  the  reality  of  the 
third  thesis  of  Behring  and  Kitasato;  we  can  pre- 
vent and  we  may  cure  tetanus  by  the  serum  of  ani- 
mals vaccinated  against  this  disease. 

Erhlich  has  contributed  other  interesting  docu- 
ments to  this  question  of  the  antitoxines.  He  has  seen 
that  if  we  vaccinate  against  one  of  the  vegetal  toxines 
pregnant  mice,  and  give  them  after  they  have  dropped 
their  young  the  young  of  non-vaccinated  mice  to 
suckle,  these  little  mice  become  in  their  turn  refrac- 
tory to  the  given  poison.* 

These  facts  are  of  the  highest  importance,  for 
they  demonstrate  that  the  immunizing  substance  is 
secreted  in  the  milk,  and  that  it  is  absorbed  by  the 
digestive  canal.  Ehrlich  has  applied  this  datum  to 
the  infectious  diseases. 


Ehrlich,  Zeitschrift  f.  Hygiene,  t.  xii,  p.  183. 


—  104  — 

He  has  been  able  to  prove  that  the  milk  secretes 
antitetanine,  and  in  that  condition,  being  ingested, 
confers  immunity  against  tetanus.  With  Brieger  he 
has  made  other  experiments  on  a  large  scale.  A 
pregnant  goat  was  by  them  vaccinated  against  teta- 
nus, according  to  the  method  of  Brieger,  Kitasato, 
and  Wasserman.  Later  on,  the  goat's  milk,  injected 
in  mice,  immunized  them  perfectly  against  tetanus. 

Behring  and  Frank  have  recently  published  some 
interesting  researches  on  the  properties  of  the  sub- 
stance which  immunizes  against  tetanus.*  They 
found  that  the  serum  of  a  vaccinated  horse  kept  for 
two  months  its  immunizing  power,  although  preserved 
(with  a  >^-per-cent.  solution  of  phenic  acid)  in  a  flask 
closed  with  a  glass  stopper,  with  no  other  precaution 
against  the  entrance  of  the  air  or  of  microbes.  They 
found  also  that  this  immunizing  power  of  serum  is 
not  abolished  by  dilution  with  distilled  water,  or  by 
heating  it  twenty-five  minutes  at  65°  C. — when  it 
begins  to  coagulate.  They  insist  also  on  the  fact 
that  to  cure  tetanus  when  declared,  it  is  necessary  to 
employ  stronger  doses  of  the  serum  than  would  be 
required  for  propyhlaxis. 

We  must  say  a  few  words,  in  conclusion,  about 
the  therapeutic  attempts  made  on  man  with  the  im- 
munizing serum.  Kitasato  was  the  first  to  attempt 


*  Behring  and  Frank,  Deutsche  Medic.  Wochenschrift, 
1892,  No.  21. 


—  105  — 

to  cure  tetanus  in  man  by  the  serum  of  the  vaccinated 
hare.  He  employed  very  feeble  doses,  and  his  at- 
tempt was  not  crowned  with  success.  Tizzoni  and 
Cattani,  on  the  contrary,  report  already  seven  cases 
of  cure  by  their  antitetanine,  prepared  with  the  serum 
of  vaccinated  dogs. 

More  recently,  this  treatment  was  attempted 
without  success  in  two  cases  of  human  tetanus  in 
Paris.*  It  is  evident  that  there  have  as  yet  been  too 
few  cases  to  warrant  speaking  for  or  against  the 
method  of  Behring  and  Kitasato.  But  the  brilliant 
results  of  experimentation  leave  no  doubt  that  success 
on  man  is  only  a  question  of  method  and  of  doses. 


*Annales  de  1'Institut  Pasteur,  1892,  No.  4. 


CHAPTER  XL 

THE  POISONS  OF  DIPHTHERIA. 


SUMMARY. —  The  Researches  of  Roux  and  Yersin;  of 
Loffler — C.  Frdnkel  Succeeds  in  Vaccinating  Animals 
against  Diphtheria — The  Researches  of  Behring 
and  His  Discovery  of  Immunization — The  Diffi- 
culties Not  Yet  Surmounted  of  Diphtheritic  Vac- 
cination. 

In  filtering  the  diphtheritic  cultures  through  the 
Chamberland  filter,  Roux  and  Yersin  obtained  toxic 
liquids.  These  filtered  cultures  injected  in  animals 
produce  the  same  series  of  symptoms  as  the  living 
diphtheria  bacillus.  In  guinea-pigs  these  filtered  cul- 
tures generally  cause  the  following  lesions:  At  the 
place  of  inoculation  is  found  an  exudative  oedema; 
the  lymphatic  glands  are  congested;  the  small  intes- 
tine, the  lungs,  and  the  suprarenal  capsules  are  en- 
gorged; and  the  pleura  contains  a  serous  effusion. 
The  systemic  intoxication  of  hares  is  characterized 
especially  by  diarrhoea  and  fatty  degeneration  of  the 
liver.  In  smaller  doses  which  kill  only  after  a  long 
time,  the  poison  of  the  filtered  cultures  may  provoke 
in  animals  the  different  paralyses  which  are  so  fre- 
quent in  diphtheria  in  man. 


—  107  — 

The  toxic  substance  is  completely  destroyed  by 
heating  for  ten  minutes  up  to  100°  C.* 

In  another  memoir,  Roux  and  Yersin  have 
studied  at  greater  length  the  properties  of  this  diph- 
theritic poison,  which  they  have  classed  among  the 
diastases  f  We  shall  not  enter  farther  into  the  discus- 
sion as  to  the  chemical  nature  of  this  poison — a  discus- 
sion which  is  set  forth  at  length  with  all  its  biblio- 
graphical details  in  Chapter  VI. 

Loftier  has  confirmed  the  existence  of  the  diph- 
theritic poison.  He  extracted  it  by  means  of  glycerin 
from  cultures  made  on  hashed  meatj 

The  resistance  of  the  diphtheria  poison  to  the  dif- 
ferent means  of  destruction  has  been  but  little  studied. 
Heat  above  60°  C.  destroys  it,  or,  as  we  believe,  mod- 
ifies it,  leaving  the  cachectizing  poison  uninjured. 
The  same  decomposition  is  obtained  by  the  action  of 
the  peptic  diastases,  such  as  trypsin  and  pepsin.  Con- 
tact with  alcohol  modifies  also  the  diphtheritic  poison. 
The  different  oxidizing  agents,  as  potassium  perman- 
ganate, destroy  it  completely.  The  reducing  agents, 
such  as  HS2,  have  no  action  on  it. 

Roux  and  Yersin  have  completely  failed  in  their 


*Roux  and  Yersin,  Annales  de  1'Inst.  Pasteur,  1888, 
No.  12. 

f  Roux  and  Yersin:  Study  of  Diphtheria  (Annales  de 
1'Inst.  Pasteur,  1889,  p.  273). 

|  Loffler,  Deutsche  Medic.  Wochenschrift,  1889,  Nos. 
5  and  6. 


—  io8  — 

attempts  to  habituate  or  to  vaccinate  animals  against 
the  diphtheritic  poison.  C.  Frankel  has  been  more 
successful.  In  heating  cultures  of  diphtheria  at  70° 
C,  he  has  succeeded  in  depriving  them  in  great  part 
of  their  toxicity,  while  not  injuring  their  vaccinal 
power.  He  has  found  that  the  heating  of  diphtheritic 
cultures  three  weeks  old  should  be  made  at  the  tem- 
perature of  from  60°  to  70°  C.  If  this  temperature  is 
exceeded,  you  obtain  no  longer  vaccinant  effects,  and 
you  even  produce  a  slow  intoxication  of  the  animals. 
With  cultures  heated  to  70°  C.,  and  inoculated  in  the 
dose  of  from  10  to  20  cubic  centimeters  under  the 
skin  of  guinea-pigs,  immunity  is  conferred  at  the  end 
of  two  weeks.* 

Shortly  after  Frankel's  communication,  appeared 
the  memoir  of  Behring  on  the  same  subject.  Behring 
gives  five  different  processes  of  vaccinating  animals 
against  diphtheria: 

1.  With  the  sterilized  cultures,  as  in  the  method 
of  Frankel. 

2.  With  diphtheritic  cultures  to  which  has  been 
added  trichloride  of  iodine. 

3.  By  the  pleuritic  exudation  of  guinea-pigs  which 
have  died  of  diphtheria. 

4.  By  the  inoculation  of  animals  with  the  virulent 
diphtheritic  bacillus  followed  by  their    treatment  by 
trichloride  of  iodine  in  subcutaneous  injection. 

*  Vide  Gamalela,  in  C.  R.  de  la  Soc.  de  Biologic,  Feb. 
20,  1892;  also  C.  Frankel,  Berlin.  Klin.  Woch.,  1890,  No  49. 


—  109  — 

5.  By  oxygenated  water,  which  has  the  property 
of  vaccinating  the  animals  against  diphtheria,  just  as 
trichloride  of  iodine  vaccinates  them  against  tetanus. 

The  great  number  of  processes  which  were  only 
touched  upon  in  the  memoir  of  Behring  made  it  plain 
that  this  author  did  not  possess  a  good  method  of  vac- 
cination against  diphtheria.*  And  his  subsequent  pub- 
lications only  went  to  confirm  this  conviction. 

Proskauer  and  Wassermann  have  attempted  to 
vaccinate  against  diphtheria  by  the  poisons  subjected 
to  heat,  but  with  constantly  negative  results. f 

Zimmer,  at  the  laboratory  of  C.  Frankel,  has  veri- 
fied the  exactitude  of  the  assertions  of  Behring.  He 
has  noted,  in  general,  that  a  more  or  less  complete 
vaccination  may  result  from  the  employment  of  these 
different  processes,  but  that  no  one  of  them  is  certain. 
With  most  of  the  processes  of  Behring  the  immunity 
of  the  animals  is  the  exception,  and  an  ultimate  death 
is  the  rule. 

Only  one  of  the  methods  of  Behring  should  be 
signally  excepted:  it  is  that  of  the  injection  in  the 
animals  of  diphtheritic  cultures  attenuated  by  the 
addition  of  trichloride  of  iodine.  This  process  has 
given  Zimmer  positive  results. J  It  is  far,  however, 


*  Behring,  Deutsche  Medic.  Woch.,  1890,  No.  50. 
f  Proskauer  and  Wassermann,  Deutsche  Med.  Woch.. 
1891,  No.  17. 

\ Zimmer,  Deutsche  Medic.  Woch.,  1892,  No.  16. 


—   no  — 

from  constituting  a  definitive  method  of  vaccination, 
for  Behring  himself  does  not  trust  it;  in  his  most  re- 
cent work  on  this  subject  he  describes  as  his  method 
of  anti-diphteritic  vaccination  certain  extremely  com- 
plicated processes.  He  injects  successively  diphthe- 
ritic cultures  heated  to  90°,  80°,  and  70°  C.;  then 
mixtures,  at  different  degrees  of  strength,  of  cultures 
with  trichloride  of  iodine.  Despite  this  long  and 
complicated  preparation,  you  will  note  in  his  experi- 
ments that  most  of  his  vaccinated  animals  die  of 
diphtheria.  Moreover,  Behring  himself  acknowledges 
that  he  does  not  possess  a  satisfactory  method  of 
vaccination. 

We  shall  see  further  on,  what  enormous  interest 
is  attached  to  this  problem  of  how  to  find  a  sure 
method.  The  principles  which  guide  Behring  in  his 
vaccination  procedures  are  the  following: 

To  give  immunity  to  receptive  animals,  and  to 
augment  immunity  in  animals  which  already  pos- 
sess it  in  a  certain  degree,  the  vaccinal  injections 
ought  to  be  followed  by  a  reaction  of  the  organism. 
This  reaction  is  general  and  local.*  Behring  does 
not  indicate  in  what  the  first  consists;  as  to  the  local 
reaction,  it  is  a  tumor  more  or  less  circumscribed.  If 
the  vaccinal  reaction  is  nil,  the  vaccinal  injection  will 


*  Behring  and  Wernicke,  Zeitschrift  fiir  Hygiene,  t. 
xii,  p.  10.  Also,  Gamaleta's  articles  in  Ann.  de  1'Inst.  Pas- 
teur, 1888,  No.  10,  and  Centralblatt  f.  Bacteriologie,  1888,  t. 
iv,  p.  161. 


Ill 


not  give  any  immunity,  or  will  not  augment  the  pre- 
existing immunity.  If  the  reaction  is  excessive,  the 
vaccinal  injection  leads  to  an  opposite  effect;  it  aug- 
ments the  receptivity  of  the  animal  to  diphtheria.  If 
the  reaction  is  moderate,  the  animals  acquire  a  cer- 
tain degree  of  immunity,  but  only  at  the  end  of  a 
certain  time — several  weeks  or  several  months.  Ac- 
cording to  this  exposition,  you  can  see  with  what 
difficulty  and  slowness  vaccination  is  made  by  the 
procedures  of  Behring. 

Consequently,  it  is  with  reason  that  the  authors 
of  a  recent  work  on  the  subject — Brieger,  Kitasato, 
and  Wassermann — declare  that  there  does  not  thus  far 
exist  any  process  which  enables  us  to  vaccinate  with 
certainty  against  diphtheria.  In  making  use  of  Wool- 
dridge's  method,  they  give,  in  their  turn,  a  new  pro- 
cedure: they  cultivate  the  diphtheritic  bacillus  in 
extract  of  thymus,  and  then  heat  this  culture  (after 
C.  Frankel)  between  60°  and  70°  C.  for  fifteen  min- 
utes. With  the  vaccine  prepared  in  this  way,  they 
have  obtained  some  positive  results.  But  this  mixed 
procedure  is  not  more  sure.* 

Although  we  do  not  yet  possess  a  satisfactory 
method  of  vaccination  against  diphtheria,  the  fact  of 
the  possibility  of  this  vaccination  is  sufficiently  estab- 
lished. 


*Breiger,  Kitasato,  and  Wassermann,  Zeitschrift  fttr 
Hygiene,  t.  xii,  p.  137.  Also,  Behring  and  Kitasato,  Deutsche 
Medic.  Wochen.,  1890,  No.  49. 


112    

The  study  of  these  vaccinated  animals  has  led 
Behring  and  Kitasato  to  results  extremely  interesting. 

Behring*  has  noted — and  this  fact  has  been  con- 
firmed by  other  observers — that  the  inoculation  of 
the  diphtheritic  bacillus  in  vaccinated  animals  is 
followed  by  the  formation  of  a  necrotic  patch  under 
which  the  bacillus  long  lives  in  a  state  of  activity.  It 
is  not,  we  perceive,  destroyed  in  the  vaccinated  organ- 
ism. It  does  not  even  seem  to  have  lost  its  virulence, 
for  its  subsequent  inoculation  in  animals  not  vaccin- 
ated provokes  in  them  typical  diphtheria.  As,  how- 
ever, it  remains  inoffensive  to  the  vaccinated  organism 
which  gives  it  harbor,  Behring  supposed  that  the  lat- 
ter possesses  the  power  of  destroying  its  poison. 

We  have  seen  in  the  history  of  the  tetanus  poison 
how  this  supposition  was  confirmed  by  Behring  and 
Kitasato. 

For  diphtheria,  as  for  tetanus,  these  authors  found 
in  the  serum  of  vaccinated  animals  certain  remarkable 
antitoxic  properties.  The  serum  of  certain  animals 
vaccinated  against  diphtheria  —  guinea-pigs,  hares, 
sheep — has  the  power  to  destroy  in  vitro  the  diph- 
theritic poison;  it  may  consequently  prevent  and  cure 
diphtheria  in  animals  in  which  it  is  injected  in  suffi- 
cient quantity. 

If  all  these  researches  have  not  yet  led  to  any 


*  Behring:  Communication  to  the  Congress  of  Hygiene. 
London,  1891. 


good  practical  result  in  the  cure  of  diphtheria  in 
man,  this  is  due  solely  to  the  difficulty  of  procuring 
the  serum  of  vaccinated  animals  in  great  quantity; 
and  this  difficulty  depends  on  the  absence  of  a  good 
method  of  vaccination.  It  is  for  this  reason  that  we 
have  here  dwelt  at  such  length  on  the  question  of 

vaccination  against  diphtheria. 
8000 


CHAPTER  XII. 

THE    POISONS   OF  CHOLERA   AND   OF  THE 
AVICIDE    VIBRIO. 


SUMMARY. — The  Researches  of  the  Author  on  the  Poi- 
sons of  Cholera  and  of  the  Avicide  Vibrio — The 
Researches  of  Hernandez  and  Bruhl,  of  Niessen 
and  Behring,  and  of  Zasslein. 

The  history  of  the  development  of  our  sources  of 
information  respecting  the  poisons  of  cholera,  shows 
in  a  typical  manner  the  struggle  of  the  different  con- 
ceptions concerning  the  chemical  nature  of  the  micro- 
bian  poisons.  In  order  to  explain  cholera,  we  have 
seen  successively  advanced  such  considerations  as 
the  products  of  the  metabolism  of  the  Indian  vibrio, 
the  diastases  which  it  secretes,  and,  lastly,  the  action 
of  the  microbe  by  its  own  substance.*  Cantani  was 
the  first  to  indicate  that  the  cholera  poison  may  be 
neither  a  diastase  nor  a  ptomaine,  but  the  cholera 
vibrio  itself.  This  he  has  not  proved,  f 

I  have  myself  found  the  two  cholera  poisons:  the 
primary  poison  and  the  modified  poison.  The  first, 

*  Gamalela:  Experimental  Researches  on  the  Poisons  of 
Cholera  (Arch,  de  Med.  Exper.,  March  i,  1892). 

t  Cantani,  Deutsche  Medic.  Wochen.,  1886,  No.  45. 


which  we  believe  to  be  a  nucleo-albumin,  has  the 
property  of  provoking  a  systemic  intoxication  ex- 
ceedingly like  the  symptoms  of  Asiatic  cholera.  In 
hares  this  intoxication  manifests  itself  chiefly  by  a 
prolonged  and  violent  diarrhoea;  in  dogs  the  vomit- 
ings predominate,  and  may  last  several  hours.  In  all 
cases  the  poison  determines  a  violent  lesion  of  the 
gastro-intestinal  tube. 

This  poison  is  extremely  fragile.  It  is  destroyed 
by  heat  above  60°  C.,  by  alcohol,  and  by  the  strong 
alkalies.  It  is  carried  down  by  the  different  precipi- 
tates which  may  be  produced  in  the  cholera  cultures; 
for  example,  by  that  which  is  formed  by  acetate  of 
lead.  It  may  be  extracted  from  them  by  alkalinized 
water. 

What  is  the  mode  of  action  of  this  poison  ?  By 
some  of  its  reactions  it  resembles  the  soluble  fer- 
ments. We  know  also  that  the  cholera  vibrio  may 
produce  the  different  diastases.  It  may  be  asked  if 
it  is  not  these  diastases  which  are  toxic  in  the  cultures 
of  cholera?  Some  think  that  the  excessive  produc- 
tion of  liquid  in  cholera  stools  is  the  result  of  a  hyper- 
secretion  of  the  intestinal  glands  (Cohnheim),  and  we 
know  that  the  substances  secreted  by  the  glands 
stimulate  their  activity  if  injected  into  the  blood.  In 
studying  the  toxic  action  of  the  cultures  of  different 
microbe  producers  of  diastases,  we  find,  in  fact,  that 
they  all  have  as  a  common  character  a  certain 
diarrhceic  action,  though  this  is  most  marked  in 


—  n6  — 

the  cholera  cultures.  Even  certain  samples  of  pan- 
creatin,  in  our  experiments  produced  diarrhoea  in 
hares.  But  the  results  with  this  latter  substance  have 
not  been  constant,  and  we  have  at  least  succeeded  in 
procuring  some  trypsin  which  is  extremely  active  as  a 
peptic  diastase  and  which  has  no  diarrhceic  action. 
Many  other  ferments  which  we  have  studied  in  this- 
connection  were  also  devoid  of  the  property  of  pro- 
voking diarrhoea. 

We  must  conclude  that  the  diarrhceic  poison  of 
cholera  does  not  belong  to  the  category  of  these  fer- 
ments. 

The  diarrhceic  action  is  very  general  in  the  class- 
of  microbian  poisons,  but  it  is  due  to  another  mechan- 
ism than  that  indicated  above. 

We  have  found  still  another  cholera  poison, 
which  is  very  stable.  It  belongs  to  the  class  of  our 
modified  poisons,  or  nucleines,  which  Buchner  has  de- 
nominated proteins  or  alkali-albumins.  This  poison 
has  no  other  interest  than  that  of  being  associated 
with  the  vaccinal  substance  of  cholera.  But  we  shall 
not  here  set  forth  the  details  of  cholera  vaccination,, 
which  is  still  under  controversy.  There  exists  a 
microbe  which  resembles  in  all  respects  that  of 
cholera.  It  is  the  avicide  vibrio,  which  I  discovered 
at  Odessa.  Chemical  vaccination  with  this  microbe 
is  at  present  the  best  known  of  all  the  vaccinations. 
As,  besides,  it  has  many  analogies  with  cholera  vac- 
cinations, we  shall  venture  to  say  a  few  words  about  it. 


The  vaccinal  substance  or  substances  which  are 
potent  against  the  avicide  vibrio  are  also  associated 
with  the  nucleine  or  modified  poison  of  the  vibrio. 
They  may  be  isolated  in  two  ways.  First,  if  you  sub- 
ject to  distillation  in  a  vacuum  the  vibrionic  cultures, 
the  substances  which  pass  over  have  a  manifest  vac- 
cinal action,  while  the  toxic  nucleine  remains  in  the 
residue.  Now  Briihl  has  found  that  if  we  precipitate 
the  toxic  substance  by  acetate  of  lead,  the  filtrate 
vaccinates  perfectly.  The  vaccinal  action  of  the 
volatile  products  would  lead  one  to  think  that  the 
vaccine  is  a  ptomaine,  but  the  ordinary  methods  for 
the  isolation  of  the  ptomaines  have  resulted  only  in 
inactive  substances  (Bruhl).  Alcohol  especially  seems 
to  enfeeble  the  vaccinal  properties  of  the  vibrionic 
cultures. 

The  study  of  the  serum  of  animals  vaccinated 
against  either  the  avicide  vibrio  or  the  cholera  vibrio 
reveals  certain  remarkable  peculiarities.  While  the 
serum  of  ordinary  guinea-pigs  favors  the  growth 
of  the  avicide  and  cholera  vibrios,  the  serum  of 
guinea-pigs  vaccinated  against  these  diseases  acquires 
the  property  of  completely  destroying  these  microbes. 
We  have  no  need  of  insisting  upon  the  bearing  of 
these  facts  on  the  true  solution  of  the  problem  of  im- 
munity. 

Still  another  fact  of  considerable  theoretical  im- 
portance has  been  found  in  connection  with  the  dis- 
eases caused  by  these  two  vibrios.  Guinea-pigs  which 


—  n8  — 

are  vaccinated  against  the  avicide  or  cholera  vibrio 
are  not  more  resistant  to  the  vaccinal  poison  than 
non-vaccinated  guinea-pigs.  This  fact  is  of  funda- 
mental interest  in  reference  to  the  entire  mechanism 
of  vaccination. 

Denied  at  first  by  certain  bacteriologists,  it  was 
later  affirmed  by  the  same,  and  adduced  in  opposition 
to  the  great  principles  established  for  tetanus  and 
diphtheria.  We  have  seen  that  in  these  two  diseases 
the  animals  which  are  vaccinated  against  the  living 
microbe  become  also  refractory  to  the  microbian 
poisons. 

Cholera  and  the  vibrionic  septicaemia  seemed  to- 
be  an  exception  to  this  rule,  for  in  these  two  diseases 
the  animals  succumb  to  the  same  dose  of  the  poison 
as  the  control  animals.  But  this  exception  is  only 
apparent,  and  it  disappears  when  we  recognize  the 
distinction  between  the  primitive  and  the  modified 
poisons  of  bacteria. 

Towards  the  primitive  poisons  of  cholera  and  of 
septicaemia,  the  vaccinated  animals  are  quite  as  re- 
fractory as  is  the  case  with  diphtheria  and  tetanus. 
They  are  not  refractory  towards  the  modified  poisons; 
but  as  these  latter  are  artificial  products,  this  has 
nothing  to  do  with  the  theories  of  immunity. 

With  regard  to  the  mechanism  of  vaccination,  it 
may  appear  strange  that  the  modified  poisons  to 
which  the  animal  is  not  habituated  render  it  refrac- 
tory to  the  primitive  poisons  and  to  the  bacteria. 


—  n9  — 

Here  also  we  must  make  a  distinction.  The  vaccinal 
substance  is  not  the  modified  poison,  for  it  may  be 
separated  from  it.  Not  being  toxic,  it  does  not  vac- 
cinate the  animals  by  the  fact  of  habituation,  but 
probably  by  forming  in  their  bodies  a  compound 
which  has  antitoxic  properties. 


r 

CHAPTER  XIII. 

THE  POISONS   OF  TUBERCULOSIS. 


SUMMARY. — The  Researches  of  Koch,  of  Maffuci,  of 
Prudden  and  Ifodenpyl,  of  Straus  and  the  Author, 
of  Grancher  and  Ledoux-Lebard — The  Tubercu- 
lin of  Koch,  and  the  Researches  which  it  has  Pro- 
voked—  The  Toxomucin  of  WeyL 

The  tuberculous  poisons  have  only  lately  been 
found.  Bacteriologists  were  long  and  unsuccessfully 
searching  for  the  toxic  substances  in  the  cultures 
of  Koch's  bacillus,  and  they  have  only  found  them  by 
applying  directly  to  the  bacillus  itself. 

Koch  has  seen  that  the  bacillus  tuberculosis 
when  dead  provokes  by  subcutaneous  injection  the 
formation  of  abscesses  in  animals.  Moreover,  he  has 
obtained  from  the  bodies  of  the  bacilli  a  substance 
called  tuberculin,  which  has  long  been  believed  to 
exercise  a  specific  action  on  the  tuberculous.* 

Maffuci  has  found  a  cachectizing  influence  in 
tuberculous  cultures  dead  by  age  or  killed  by  dis- 
continuous sterilization.f 


*  Koch,    Deutsche   Medic.  Wochenschrift,  Jan.  18,  1891. 
f  Maffuci,  Centralblatt  f.  Allgemeine  Pathol.,  Dec.  15, 
1890. 


121    

Prudden  and  Hodenpyl  have  seen  that  the  bacilli 
when  boiled  and  well  washed  may  give  rise  to  the 
formation  of  curable  nodular  lesions.* 

Straus  and  I  have  shown  that  the  cadavers  of 
tubercle  bacilli  provoke  in  different  animals  a  disease 
which  in  its  course  and  its  lesions  is  extremely  like 
tuberbulosis  produced  by  living  bacilli.  This  disease, 
which  Grancher  and  Ledoux-Lebard  have  propossd 
to  call  necro-tuberculosis,f  is  characterized  by  the 
•development  in  inoculated  animals  of  a  great  sensi- 
tiveness towards  a  new  inoculation  of  tuberculosis. 
This  predisposition  manifests  itself  by  rapid  death 
after  re-inoculation.  The  inoculation  of  dead  bacilli 
produces  such  changes  in  the  animal  organism  that 
the  animals  die  very  speedily  after  a  new  injection  of 
tuberculosis,  an  injection  which  produces  only  tardy 
effects  in  healthy  animals. 

Another  symptom  of  necrotuberculosis,  but  much 
more  apparent,  is  the  continuous  and  progressive 
emaciation  of  the  animals.  They  may  lose  as  much 
as  one-half  of  their  initial  weight.  They  die  in  a  pro- 
found cachexia.  At  the  autopsy  we  find  characteristic 
lesions. 

The  different  organs,  and  especially  the  lungs,  in 


*  Prudden  and  Hodenpyl,   New  York  Medical  Journal, 
June  6  and  June  20,  1891. 

.  f  Straus  and  Gamalela,  Archives  de  Medecine  Experi- 
tnentale,  1891,  No.  6.  Grancher  and  Ledoux-Lebard,  Arch, 
de  M6d.  Exper.,  1892,  No.  I. 


122    

the  case  of  intravenous  injection  of  dead  bacilli,  are 
strewn  with  granulations  formed  of  epithelioid  and 
embryonal  cells,  and  containing  tubercle  bacilli.  The 
volume  of  the  nodules  which  may  be  thus  produced 
in  the  different  organs  depends  almost  exclusively  on 
the  size  of  the  mass  of  cadavers  which  has  been  inoc- 
ulated. It  is  very  interesting  to  note  that  the  animal 
organism  cannot  dislocate  the  bacterial  colonies  which 
are  inoculated  into  it. 

When  once  the  existence  of  the  tuberculous  poi- 
son, and  its  presence  in  the  cadavers  of  the  bacilli, 
was  established,  it  was  necessary  to  seek  to  determine 
the  toxic  substance  and  to  isolate  it  from  the  bacilli. 

There  were  evidently  different  means  for  pro- 
ceeding to  this  extraction.  By  alcohol  and  ether 
Hammerschlag  has  extracted  from  the  bacilli,  in  addi- 
tion to  lecithin  and  fat,  a  toxic  substance  which  kills- 
animals  with  convulsions.*  Later,  Zuelzer  also  suc- 
ceeded in  finding  a  convulsivant  ptomaine  in  tubercu- 
lous cultures.f 

Much  more  interesting  was  Koch's  discovery. 
By  means  of  glycerin-water  at  a  boiling  temperature 
and  in  a  neutral  medium,  Koch  has  extracted  from 
tubercle  bacilli  an  albuminoid  substance  called  tuber- 
culin, which  provokes  in  tuberculous  animals  fever 

*  Hammerschlag,  Correspondenzblatt  fur  Schweizer- 
Srzte,  Oct.  15,  1888,  and  Centralblatt  f.  Klin.  Med.,  Jan.  I, 
1891- 

f  Zuelzer,  Berlin.  Klin.  Wochen.,  Jan.  26,  1891. 


—  123  — 

and  inflammation  of  the  bacillary  foci.  Koch  has 
studied  in  detail  the  many  very  interesting  properties 
of  this  tuberculin.*  Tuberculin  dissolved  in  glycerin- 
water  is  a  very  stable  body  which  supports  a  heat  of 
1 60°  C.  It  is  easily  precipitable  by  alcohol. 

But  if  we  attempt  to  purify  it  and  to  free  it  from 
glycerin  and  salts,  its  properties  are  modified.  Aque- 
ous solutions  of  purified  tuberculin  are  very  unstable 
in  absence  of  glycerin.  They  easily  give  a  precipitate 
more  or  less  abundant  of  a  substance,  insoluble  in 
water  but  soluble  in  alkalies,  which  preserves  the  toxic 
action  of  the  tuberculin.  In  absence  of  salts,  the 
purified  tuberculin  is  no  longer  precipitated  by  abso- 
lute alcohol. 

The  purified  tuberculin  has  all  the  reactions  of 
albuminoid  substances.  It  gives  a  precipitate  by 
acetic  acid  which  is  soluble  in  excess  of  the  reagent. 
It  contains  a  great  quantity  of  phosphorus.  But  it  is 
evidently  not  a  pure  substance,  for  it  leaves  from  14 
to  21  per  cent,  of  ashes,  and  then  its  mode  of  prepa- 
ration by  the  concentration  of  cultures  made  in 
bouillon  adds  to  the  extract  of  bacillary  bodies  all 
the  substances  pre-existing  in  the  bouillon. 

Hunter  f  has  endeavored  to  advance  still  farther 
our  knowledge  of  tuberculin.  Unfortunately  he  also 


*  Koch,  Deutsche  Medic,  Wochen.,  1891,  No.  43.     Gama- 
lela,  Arch,  de  Med.  Exper.,  1891,  No.  2. 

f  British  Medical  Journal,  July  28,  1891. 


—   124  ~ 

made  use,  in  his  chemical  researches,  of  the  crude 
tuberculin  of  Koch,  prepared  by  means  of  culture 
bouillons,  instead  of  taking  the  bacilli  themselves, 
washed  and  cleaned.  In  this  crude  tuberculin  Hunter 
found  the  four  albumoses  of  Kuehne  and  Chittenden. 
It  is  to  be  regretted  that  he  did  not  attempt  to  find 
them  in  the  primitive  bouillon  of  the  cultures. 

Hunter  has  also  compared  the  differences  in  the 
action  on  the  tuberculous  man  and  animal  of  the  dif- 
ferent modified  tuberculins.  The  modification  A, 
which  is  the  precipitate  obtained  by  absolute  alcohol, 
has  the  property  of  producing  the  local  reaction  with 
very  little  fever.  The  modification  C,  which  contains 
the  substances  not  precipitated  by  absolute  alcohol, 
produces  chiefly  a  high  fever.  The  modification  B, 
which  is  the  precipitate  obtained  by  supersaturation 
of  the  tuberculin  with  ammonium  sulphate,  possesses 
the  power  of  provoking  the  local  and  salutary  reac- 
tion without  constitutional  disorders.  Lastly,  the 
modification  CB  has  all  the  curative  virtues  of  Koch's 
tuberculin,  without  possessing  the  disadvantages — 
either  the  local  inflammatory  or  the  general  febrile 
reaction.  This  happy  modification,  CB,  is  obtained 
by  getting  rid,  by  absolute  alcohol,  of  most  of  the 
albumins,  and  by  the  dialysis  of  other  injurious  sub- 
stances. These  results  of  Hunter  are  founded  on  an 
altogether  insufficient  number  of  experiments.  Fur- 
thermore, according  to  all  the  other  experimenters, 
and  particularly  Pfuhl,  the  collaborator  of  Koch,  tuber- 


—  125  — 

culin  has  not  any  curative  properties,  especially  if  you 
prevent  the  local  reactions.* 

The  tuberculocidin  of  Klebs,  prepared  by  means 
of  tuberculin,  is  entitled  to  no  greater  confidence. 

As  to  the  chemical  nature  of  tuberculin,  we  can- 
not pronounce  with  certainty  until  we  have  a  purer 
preparation.  There  is  no  doubt  that  it  belongs  to 
our  class  of  modified  poisons,  which  we  suppose  to  be 
nucleines  or  nucleinic  acids.  It  is  evident,  also,  that 
in  Koch's  tuberculin  is  not  found  the  primary  tuber- 
culous poison.  The  action  of  tuberculin  does  not 
explain  the  effects  produced  by  the  dead  bacilli. 
This  action  is  not  characteristic  of  tuberculin,  for  it 
is  also  provoked  by  many  other  bacterian  extracts. 
The  effects  proper  to  the  tuberculous  poison  con- 
tained in  the  microbian  cadavers,  the  predisposition 
which  it  creates,  and  the  suppurative  and  caseous 
lesions,  cannot  be  produced  by  tuberculin. 

More  important  in  this  respect  is  the  substance 
which  Weyl  has  extracted  from  tubercle  bacilli  by 
means  of  a  solution  of  caustic  soda.  This  substance, 
insoluble  in  acetic  acid,  is  classed  by  Weyl  among  the 
mucins,  but  it  does  not  form  a  reducing  substance, 
and  it  contains  phosphorus.  It  may  be  nucleo-albu- 
min  in  the  sense  of  Kossel  and  Hammersten. 

This  mucin  of  Weyl  has  a  toxic  action.  Bjr 
subcutaneous  injection  it  produces  in  mice  and 


*  Pfuhl,  Zeitschrift  ftir  Hygiene,  t.  xi,  page  241. 


—    126    — 

guinea-pigs  a  necrosis  of  the  skin.  But  the  toxi- 
cological  study  of  Weyl  is  too  incomplete.  We  cannot 
but  suppose  that  he  had  under  observation  the  actual 
primary  tuberculous  poison,  that  which  produces  the 
local  lesions  of  tuberculosis.* 


*Weyl,  Deutsche  Medic.  Wochen.,  Jan.  12,  1891. 


CHAPTER  XIV. 

THE  POISONS  OF  CHARBON  AND  OFGLANDERS. 


SUMMARY. — Insufficiency  of  our  Toxicological  Knowl- 
edge respecting  Charbon  and  Glanders — Different 
Writings  on  the  Subject. 

Charbon  (splenic  fever,  wool-sorters'  disease, 
deadly  anthrax),  which  once  had  so  great  a  doctrinal 
importance  as  to  guide  the  first  steps  of  students  in 
bacteriology,  has  not  preserved  this  eminent  place 
in  toxicological  researches. 

Toussaint  believed  that  he  could  confer  immu- 
nity on  sheep  and  young  dogs  by  means  of  anthracoid 
blood  freed  from  living  bacteria  by  filtration  or  by 
heating  to  55°  C.,  and  even  by  antiseptics. 

Chauveau  brought  powerful  arguments  in  favor 
of  this  chemical  vaccination.* 

After  having,  with  Pasteur,  denied  its  reality, 
Chamberland  and  Roux  published  later  some  experi- 
ments which  showed  that  it  is  in  fact  possible  to  vac- 
cinate sheep  against  charbon  by  means  of  soluble 
substances,  f 


*Annales  de  1'Institut  Pasteur,  1888,  No.  2. 
\  Chamberland  and  .Roux,  Ann.    de  1'Institut  Pasteur, 
1888,  No.  7. 


—    128    — 

Already,  moreover,  before  these  experiments,  this 
possibility  had  been  demonstrated  by  Wooldridge. 
He  found  that  anthracoid  cultures  made  in  a  solution 
of  the  substance  which  he  called  tissue-fibrinogen* 
might  confer  immunity  on  hares  if  injected  subcuta- 
neously,  when  freed  from  living  microbes  by  ebullition 
or  by  filtration.  What  is  surprising  in  the  experi- 
ments of  Wooldridge  is  that  his  substance  conferred 
immunity  not  only  when  it  was  injected  long  before 
the  injection  of  the  living  and  virulent  microbe,  but 
also  in  the  case  of  its  simultaneous  injection  with  the 
microbe.  Wooldridge  thus  furnishes  the  first  in- 
stance of  the  experimental  cure  of  an  infection  by 
means  of  vaccination. f 

The  interesting  researches  of  Wooldridge,  inter- 
rupted by  the  premature  death  of  this  young  savant, 
have  only  recently  been  taken  up  again  by  Brieger,. 
Kitasato,  and  Wassermann.  But  what  is  a  little 
singular,  the  authors  having  very  well  succeeded  in 
preparing  by  the  method  of  Wooldridge  chemical 
vaccines  against  the  different  pathogenic  microbes, 
have  been  less  fortunate  with  charbon.  Perhaps 


*  Wooldridge,  Arch.  f.  Anatomic  und  Physiologic, 
Physiologische  Abtheilung,  1888,  p.  527.  Wooldridge's  first 
publications  date  from  1887. 

fThe  fibrinogenous  substances  of  Wooldridge  are  com- 
pounds between  the  albuminoids  and  lecithin.  He  extracted 
them  from  different  fresh  organs,  and  especially  from  the 
thymus  and  testicles  of  animals. 


—    I29    — 

these  men  have  not  been  sufficiently  persevering  in 
this  direction,  having  the  old  prejudice  as  to  the 
radical  distinction  between  the  toxic  diseases  (tetanus, 
cholera,  diphtheria,  typhoid  fever)  on  the  one  hand, 
and  the  septicaemias  (charbon,  rouget  of  swine,  etc.) 
on  the  other.* 

Wooldridge's  experiments  on  charbon  have  also 
been  continued  by  Hankin  and  Martin. f 

Hankin  has  cultivated  the  anthrax  bacillus  in 
Liebig's  beef-extract  with  the  addition  of  fibrin. 
From  these  cultures  he  has  isolated,  by  precipitating 
them  with  alcohol,  an  albuminoid  substance  which  he 
calls  albumose.  This  albumose  has  the  property  of 
vaccinating  against  charbon,  and  even  of  curing  this 
disease. 

We  must,  however,  immediately  add  that  Peter- 
mann,  who  has  repeated  exactly  the  experiments  of 
Hankin,  has  arrived  at  entirely  negative  results.  Be- 
fore Petermann,  Landi  also  obtained  only  negative 
results.  Martin|  has  cultivated  the  anthrax  bacillus  in 
a  solution  of  pure  alkali-albumin.  He  has  studied  in 
detail  all  the  chemical  products  which  form  in  cul- 


*Brieger,  Kitasato,  and  Wassermann,  Zeitschrift  f. 
Hygiene,  t.  xii,  fasc.  2. 

f  Hankin,  British  Medical  Journal,  Oct.  12,  1889,  p. 
810. 

$  Sidney  Martin,  Proceedings  of  the  Royal  Society, 
May,  1890.  Annual  Report  of  the  Local  Government  Board, 

London,  1889-1890,  p.  235. 
9000 


—  130  — 

ture-broths.  In  accord  with  Kuehne,  he  has  found 
the  three  albumoses  and  the  peptone  which  result 
from  the  digestion  of  albuminoids  by  the  proteolytic 
ferments.  He  has  also  found  an  organic  base.  The 
albumoses  and  the  base  are  all  toxic,  especially  the 
latter.  A  toxic  ptomaine  was  long  since  isolated 
from  anthracoid  cultures  by  Hoffa,*  who  employed 
three  different  methods  for  its  extraction,  and  who 
cultivated  the  anthrax  bacterium  on  meat,  or  on  eggs 
mixed  with  bouillon.  The  ptomaine  of  Hoffa  is, 
however,  very  little  toxic  like  that  of  Martin. 

Lando  Landi  f  has  made  some  very  interesting 
researches  on  the  substances  produced  by  the  anthra- 
coid bacterium.  He  has  found  abase  toxic  to  mice,  of 
the  carbopyridic  series,  and  some  peculiar  albumoses, 
which  may  be  obtained  in  a  crystalline  state;  but  his 
attempts  to  vaccinate  animals  with  all  these  products 
have  not  succeeded. 

Lastly,  Christmas}  seems  to  have  obtained  some 
positive  results.  He  vaccinates  hares  against  char- 
bon  by  two  processes:  First,  by  the  diseased  anthra- 
coid organs  crushed  and  emulsified  in  water,  in  which 
the  bacteria  are  killed  by  eucalyptus  essence,  and 
the  products  then  filtered  through  paper.  In  his 


*  Hoffa:  Die  Natur  des  Melzbrand  Giftes.  Wiesbaden, 
1886. 

f  Lando  Landi,  C.  R.  de  la  Soc.  de  Biologic,  July  25, 
1892. 

:{:  Christmas,  Ann   de  1'  Inst.  Pasteur,  1891,  p.  487. 


second  process  he  makes  anthrax  cultures  in  a 
medium  composed  of  yolks  of  eggs,  egg  albumin, 
and  veal  broth  feebly  alkaline,  in  equal  parts.  The 
bacilli  grow  very  well  in  this  medium,  but  do  not  form 
spores.  After  five  or  six  days  of  incubation  at  30° 
C  ,  the  cultures  are  diluted  with  water  and  passed 
through  the  Chamberland  filter.  The  filtered  liquid 
possesses  feeble  vaccinal  properties.  If  the  cultures 
are  prolonged  beyond  sixty-seven  days,  they  become 
very  toxic  and  not  at  all  vaccinant.  These  anthra- 
coid  poisons  have  not  been  studied  by  Christmas. 

To  sum  up,  we  see  that  despite  a  considerable 
number  of  researches  the  anthracoid  poisons  still 
remain  almost  entirely  unknown  to  us,  and  the  little 
that  we  do  know  is  not  yet  established  with  certainty. 

The  poisons  of  glanders  were  first  studied  by 
Finger.*  He  found  that  boiled  cultures  of  the 
glanders  bacillus  are  toxic;  but  the  phenomena  which 
they  produce  have  no  similarity  to  the  symptoms  of 
farcy- infection.  He  has  succeeded  in  some  excep- 
tional cases  in  conferring  immunity  on  hares  against 
the  living  microbe  by  means  of  these  boiled  cultures. 
Later,  this  question  was  again  taken  up  by  Bromberg,f 
who  has  seen  that  even  at  120°  C.  the  toxic  properties 


*  Finger,  Ziegler's  Beitrage  zur  Path.  Anatom.,  1889,  t. 
vi,  p.  373- 

f  Bromberg,  C.  R.  des  Travaux  de  1'Institut  Veterinaire 
de  Kharkow,  t.  iii,  i88g. 


—  132  — 

of  the  cultures  of  the  glanders  bacilli  are  not  com- 
pletely destroyed. 

Since  then,  the  poisons  of  glanders  have  been 
only  studied  from  the  special  point  of  view  of  malleine 
applied  to  the  diagnosis  of  latent  glanders,  as  we  have 
before  seen  (see  page  78). 

It  is  thus  seen  that  our  sources  of  knowledge  re- 
specting the  poisons  of  glanders  are  scarit. 


CHAPTER  XV. 

PRACTICAL  RESULTS  OBTAINED  BY  THE  STUDY 
OF  MICROBIAN  TOXICOLOGY. 


SUMMARY. — Rapid  Indications  respecting  the  Other  Bac- 
terial Poisons — Immunization  as  a  Result  of  Toxi- 
cological  Studies. 

We  are  obliged  to  stop  here  in  our  study  of  the 
microbe  posons.  We  must,  however,  before  closing, 
enumerate  some  of  the  special  treatises  on  the  sub- 
ject which  the  student  would  do  well  to  consult. 

First,  the  memoirs  of  Charrin  and  of  Bouchard's 
laboratory  on  the  pyocyanic  poisons;*  those  of  Mon- 
fredi  and  Traversal  and  Roger  on  the  poisons  of 
erysipelas;  J  of  Chamberland  and  Roux  on  the  septic 
poison  and  that  of  symptomatic  charbon;  §  those  of 
Arloing  on  the  poison  of  the  bacillus  heminecrobio- 


*  Charrin:  The  Pyocyanic  Disease.  Paris,  1889.  Also 
his  opening  article  in  the  new  Traite  de  Medecine,  tome  i. 

f  Monfredi  and  Traversa,  in  Giorn.  Intern.  Science 
Mediche.  1888. 

\  Roger,  C.  R.  de  la  Soc.  de  Biol.,  July  4,  i8qr. 

§  Chamberland  and  Roux,  Ann.  de  1'Institut  Pasteur, 
1887,  No.  12.  Roux,  Ann.  de  1'Institut  Pasteur.  1888,  No  2. 


-   134  — 

phylus;  *  those  of  Selander  on  hog-cholera;  f  those 
of  Hericourt  and  Rechit  and  Courmont  and  Dor  on 
the  bacillus  of  the  tuberculosis  of  birds.];  We  must 
also  cite  the  researches  of  Schiff  and  of  his  successors 
who  have  endeavored  to  solve  the  problem  proposed 
by  Stich  (see  page  5).§  But  we  hasten  to  abridge- 
all  these  details,  which  are  very  insignificant  in  com- 
parison with  the  great  principle  of  which  it  remains. 
for  us  to  to  speak.  We  refer  to  immunization. 

Thanks  to  the  impetus  which  has  been  given  the 
last  few  years  to  the  researches  of  microbian  toxicol- 
ogy, science  has  been  able  to  discover  and  establish 
definitively  on  solid  bases  a  principle  which  is  likely 
to  be  of  inestimable  value  in  medicine. 

According  to  the  doctrine  of  immunization,  it  is 
in  our  power  to  prevent  and  cure  infectious  diseases 
by  the  serum  of  animals  refractory  and  prepared.  |[ 
We  have  already  seen  how  immunization  is  realized  in 


*  Arloing:  Les  Virus.     Paris,  1891. 

f  Selander:  Contribution  -to  the  Study  of  Swine-pest 
(Ann.  de  1'Institut  Pasteur,  1890,  No.  9). 

t  Hericourt  and  Richet,  C.  R.  de  la  Soc.  de  Biol.,  1891* 
p.  470.  Courmont  and  Dor,  Arch,  de  Med.  Exper.,  1891, 
No.  6. 

§  Bouchard:  Lemons  sur  les  Auto-intoxications.  Paris,. 
1879.  Roger:  The  Action  of  the  Liver  on  Poisons.  Paris,  1886. 

||  This  preparation,  we  may  remind  the  reader,  is  effected 
by  saturating  these  refractory  animals  with  the  toxic  soluble 
or  vaccinant  products  of  the  corresponding  pathogenic  mi- 
crobes. 


the  case  of  tetanus.  It  has  been  applied  to  yet  many 
other  diseases.  First  it  has  been  established  for  fibrin- 
ous  pneumonia  by  the  labors  of  Emmerich  and  Fovit- 
sky  and  the  two  Klemperers.*  It  has  been  confirmed 
by  the  researches  of  Archaroff.f  Klemperer  has  given 
the  results  of  the  application  of  immunization  in  the 
treatment  of  forty  cases  of  pneumonia  in  man.;);  Im- 
munization has  also  been  applied  to  the  rouget  of  hogs, 
to  the  septicaemia  of  mice,  to  the  disease  caused  by 
Friedlander's  microbe,  and  the  pyocyanic  disease.§ 
More  recently  still,  a  way  has  been  found  for  giving 
immunity  against  typhoid  fever,  by  Brieger,  Kitasato, 
and  Wassermann,|  and  for  robine  by  Ehrlich.^f  Seeing 
the  great  number  of  immunizations  which  have  been 
discovered  in  the  short  space  of  about  eighteen  months, 
one  might  almost  be  warranted  in  concluding  that 
immunization  is  a  general  principle,  and  that,  conse- 
quently, all  the  infectious  diseases  are  curable  by  this 
method.  On  the  other  hand,  the  researches  of  Ehr- 


*  Emmerich  and  Fovitsky,  Mtlnch.  Medic.  Woch. ,  August 
11,  1891.  G.  and  F.  Klemperer,  Berlin.  Klin.  Woch.,  August 
24  and  31,  1891. 

fArch.  de  Med.  Exper.,  1892,  No.  4. 

j  Berlin.  Klin.  Woch.,  May,  1892. 

§  Klemperer,  Berlin.  Klin.  Woch.,  May,  1892.  See  also 
Munch.  Medic.  Woch.,  1892,  Nos.  5  and  6. 

||  Brieger,  Kitasato,  and  Wassermann,  Zeitsch.  f.  Hy- 
giene, t.  xii,  fasc.  2. 

1  P.  Ehrlich,  loc.  cit. 


-  136  - 

lich,  who  applies  immunization  against  the  poisons  of 
plants,  as  abrine,  ricine,  and  robine,  demonstrate 
peremptorily  that  it  is  against  the  soluble  poisons 
principally  that  immunization  acts,  and  that  by  neu- 
tralizing the  soluble  poison  one  becomes  master  of 
the  infection.  And  this  gives  support  still  further 
to  the  principal  thesis  of  my  book:  that  the  patho- 
genic microbes  are  injurious  only  by  their  poisons — 
that  infection  is  only  intoxication  by  the  bacterial 
poisons. 

Moreover,  it  was  in  the  experiments  with  the 
poisons  of  tetanus  and  diphtheria  that  immunization 
was  discovered.  And  it  seems  a  little  strange  that 
the  discovery  was  not  sooner  made,  for  the  studies  on 
the  microbian  poisons  are  less  complex  than  the  re- 
searches on  living  bacteria.  Thanks  to  the  relative 
simplicity  of  its  problems,  microbian  toxicology  has 
been  able  to  free  itself  of  the  first  of  the  clogs  of 
preconceived  ideas  which  impede  and  obstruct  the 
other  domains  of  bacteriology. 


Antiseptics  and  Disinfectants 

We  desire  to  call  attention  to  the  following  antiseptic  and  disinfectant 
preparations: 

Ethereal  Antiseptic  Soap  (Johnston's)  is  a  hydro-alcoholic  solution 
of  Castile  soap  which  was  devised  by  an  experienced  nurse  in  the  surgi- 
cal clinic  of  the  Jefferson  Medical  College.  It  may  be  made  weak  or 
strong  in  antiseptic  value  by  dissolving  mercuric  chloride  in  it  in  propor- 
tions indicated  in  the  case  in  hand.  Since  its  introduction  its  use  has 
been  extended  to  the  treatment  of  parasitic  affections  with  much  success. 

Antiseptic  Liquid  arrests  decomposition  and  destroys  noxious 
gases  that  emanate  from  organic  matter  in  sewers  and  elsewhere,  and 
may  be  used  in  cellars,  barns,  outhouses,  and  the  sick-room.  (Send  for 
Note  01  the  Disinfectant  of  the  Future,  by  Prof.  Alfred  L.  Loomis  ) 

Antiseptic  Tablets  are  convenient  for  the  extemporaneous  prepa- 
ration of  antiseptic  solutions  of  definite  strength  for  disinfectant  purposes 
and  for  antiseptic  sprays. 

Eucalyptus  and  Thymol  Antiseptic  is  adapted  for  use  as  an  anti- 
septic internally,  externally,  hypodermatically,  as  a  douche,  a  spray,  by 
atomization,  and  as  a  deodorant.  Its  application  in  surgery  is  unlimited. 
It  combines  the  antiseptic  virtues  of  benzoic  acid,  boric  acid,  oil  pepper- 
mint, oil  eucalyptus,  oil  wintergreen,  oil  thyme  and  thymol. 

Disinfectant  Powder  possesses  in  a  high  degree  disinfectant,  ab- 
sorbent, and  antiseptic  properties.  It  is  admirably  adapted  for  the  dis- 
infection of  excreta  in  cholera,  yellow  fever,  and  typhoid  fever,  and  in 
all  diseases  in  which  such  an  agent  is  indicated  for  the  purpose  specified. 

Labarraque's  Solution  we  supply  for  the  use  of  those  who  desire 
to  employ  it  for  its  local  or  internal  antiseptic  action.  It  may  be  diluted 
to  suit  the  indications. 

Sulphur  Candles  thoroughly  employed  are  effectual  in  the  fumiga- 
tion and  disinfecting  of  rooms  after  infectious  diseases. 


We  shall  be  pleased  to  forward,  on  request,  any  information  desired  con- 
cerning these  products. 


PARKE,  DAVIS  &  CO., 

Detroit,   New  York,   Kansas   City,  and  Walkerville,  Ont. 


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SERIES  I. 


Inhalers,  Inhalations  and  Inhalants. 

By  Beverley  Robinson,  M.  D. 
The  Use  of  Electricity  in  the  Removal  of 
Superfluous  Hair  and  the  Treatment  of 
Various  Facial  Blemishes. 

By  Geo.  Henry  Fox,  M.  D. 
New  Medications,  Vol.  I. 

By  Dujardin-Beaumetz,  M.  D. 
New  Medications,  Vol.  II. 

By  Dujardin-Beaumetz,  M.  D. 
The  Modern  Treatment  of  Ear  Diseases. 

By  Samuel  Sexton,  M.  D. 
The  Modern  Treatment  of  Eczema. 

By  Henry  G.  Piffard,'M.  D. 


Antiseptic  Midwifery. 

By  Henry  J.  Garrigues,  M.  D. 
On  the  Determination  of  the  Necessity  for 
Wearing  Glasses. 

By  D.  B.  St.  John  Roosa,  M.  D. 
The  Physiological, Pathological  and  Ther- 
apeutic Effects  of  Compressed  Air. 

By  Andrew  H.  Smith,  M.  D. 
GranularLids  and  ContagiousOphthalmia. 

By  W.  F.  Mittendorf,  M.D. 
Practical  Bacteriology. 

By  Thomas  E.  Satterthwaite,  M   D . 
Pregnancy,    Parturition,    the    Puerperal 
State,  and  their  Complications. 

By  Paul  F.  Munde\  M.  D. 


SERIES  II. 


The  Diagnosis  and  Treatment  of  Haem- 
orrhoids* 

By  Chas.  B.  Kelsey,  M.  D. 
Diseases  of  the  Heart,  Vol.  I. 

By  Dujardin-Beaumetz,  M.  D. 
Diseases  of  the  Heart,  Vol.  II. 

By  Dujardin-Beaumetz,  M.  D. 
The  Modern  Treatment  of  Diarrhoea  and 
Dysentery. 

By  A.  B.  Palmer,  M.  D. 
Intestinal  Diseases  of  Children,  Vol.  I. 

By  A.  Jacobi,  M.  D. 
Intestinal  Diseases  of  Children,  Vol.  II. 

By  A.  Jacobi,  M.  D. 


The  Modern  Treatment  of  Headaches. 
By  Allan  McLane  Hamilton,  M.  D. 

The  Modern  Treatment  of   Pleurisy  and 
Pneumonia. 

By  G.  M.  Garland,  M.  D. 

Diseases  of  the  Male  Urethra. 

By  Fessenden  N.  Otis,  M.  D. 
The  Disorders  of  Menstruation. 

By  Edward  W.  Jenks,  M.  D. 

The  Infectious  Diseases,  Vol.  I. 
By  Karl  Liebermeister. 

The  Infectious  Diseases,  Vol.  II. 
By  Karl  Liebermeister. 


SERIES   III. 


Abdominal  Surgery. 

By  Hal  C.  Wyman,  M.  D. 
Diseases  of  the  Liver. 

By  Dujardin-Beaumetz,  M.  D. 
Hysteria  and  Epilepsy. 

By  J.  Leonard  Corning,  M.  D. 

Diseases  of  the  Kidney. 

By  Dujardin-Beaumetz,  M.  D. 

The  Theory  and  Practice  of  the  Ophthal- 
moscope. 

By  J.  Herbert  Claiborne,  Jr.,  M.  D. 

Modern  Treatment  of  Bright's  Disease. 
By  Alfred  L.  Loomis,  M.  D. 


Clinical  Lectures  on  Certain  Diseases  of 
the  Nervous  System. 

By  Prof.  J.  M.  Charcot,  M.  D. 

The  Radical  Cure  of  Hernia. 

By  Henry  O.  Marcy,  A.  M.,  M.  D., 

LL.  D. 
Spinal  Irritation. 

By  William  A.  Hammond,  M.  D. 
Dyspepsia. 

By  Frank  Woodbury,  M.  D. 
The  Treatment  of  the  Morphia  Habit. 

By  Erlenmeyer. 

The  Etiology,  Diagnosis  and  Therapy  of 
Tuberculosis 

By  Prof.  H.  von  Ziemssen. 


SERIES 


Nervous  Syphilis. 

ByH.  C.  Wood,  M.  D. 
Education  and   Culture  as  correlated  to 
the  Health  and  Diseases  of  Women. 

By  A.  J.  C.  Skene,  M.  D. 
Diabetes. 

By  A.  H.  Smith,  M.  D. 
A  Treatise  on  Fractures. 

By  Armand  Despres,  M.  D. 
Some  Major  and  Minor  Fallacies  concern- 
ing Syphilis. 

By  E.  L.  Keyes,  M.  D. 
Hypodermic  Medication. 

By  Bourneville  and  Bricon. 


Practical    Points   in  the  Management  of 
Diseases  of  Children. 

By  I.  N.  Love,  M.  D. 
Neuralgia. 

By  E.  P.  Hurd,  M.  D. 
Rheumatism  and  Gout. 

By  F.  Le  Roy  Satterlee,  M.  D. 
Electricity,  Its  Application  in  Med'cme. 

By  Wellington  Adams,  M.D.    [Vol.1.] 
Electricity,  Its  Application  in  Medicine. 

By  Wellington  Adams,  M.D.  [Vol.11.] 
Auscultation  and  Percussion. 

By  Frederick  C.  Shattuck,  M.  D. 


SERIES   V. 


Taking  Cold. 

By  F.  H.  Bosworth,  M.  D. 

Practical  Notes  on  Urinary  Analysis. 
By  William  B.  Canfield,  M.  D. 

Practical  Intestinal  Surgery.    Vol.  I 

By  F.  B.  Robinson,  M.  D. 
Practical  Intestinal  Surgery.    Vol.  II. 

By  F.  B.  Robinson,  M.  D. 

Lectures  on  Tumors. 

By  John  B.  Hamilton,  M.  D.,  LL.  D. 

Pulmonary  Consumption,  a  Nervous  Dis- 
ease. 

By  Thomas  J.  Mays,  M.D. 


Artificial  Anaesthetics  and  Anaesthesia. 
By  DeForest  Willard,  M.  D.,  and  Dr. 
Lewis  H.  Adler,  Jr. 

Lessons  in  the  Diagnosis  and  Treatment 
of  Eye  Diseases. 

By  Casey  A.  Wood,  M.  D. 
The  Modern  Treatment  of  Hip  Disease 

By  Charles  F.  Stillman.  M.  D. 
Diseases  of  the  Bladder  and  Prostate. 

By  Hal  C.  Wyman.  M.  D. 
Cancer. 

By  Daniel  Lewis,  M.  D. 
Insomnia  and  Hypnotics. 

By  Germain  See. 

[Translated  by  E.  P.  Hurd,  M.  D.] 


The  Uses  of  Water  in  Modern  Medicine. 
By  Simon  Baruch,  M.  D.        Vol.  I . 

The  Uses  of  Water  in  Modern  Medicine, 
By  Simon  Baruch,  M.  D.      Vol.  II. 

The  Electro-Therapeutics  of  Gynaecol- 
ogy. Vol.  I. 
By  A.  H.  Goelet,  M   D. 

The  Electro-Therapeutics  of  Gynaecol- 
ogy. Vol.  II. 
By  A.  H.  Goelet,  M.  D. 


Cerebral  Meningitis. 

By  Martin  W.  Barr,  M.  D. 

Contributions  of  Physicians  to   English 
and  American  Literature. 
By  Robert  C.  Kenner,  M.  D. 


SERIES   VI. 

Gonorrhoea  and  Its  Treatment. 
By  G.  Frank  Lydston,  M.  D. 

Acne  and  Alopecia. 

By  L.  Duncan  Bulkley,  M.  D. 

Fissure  of  the  Anus  and  Fistula  in  Ano. 
By  Dr.  Lewis  H.  Adler,  Jr. 

The  Surgical  Anatomy  and   Surgery  of 
the  Ear. 
By  Albert  H.  Tuttle,  M.  D.,  S.  B. 

Recent  Developments  in  Massage. 
By  Douglas  Graham,  M.  D. 

Sexual  Weakness  and  Impotence. 
By  Edward  Martin,  M.  D. 


SERIES  VII.* 


Appendicitis  and  Perityphlitis. 

By  Charles  Talamon,  M.  D. 
Cholera.   Voi.i. 
Cholera.   Vol.  II. 

ByG.  Archie Stockwell,  M.D..F.Z.S. 
Electro-Therapeutics  of  Neurasthenia. 

By  W.  F.  Robinson,  M.  D. 
Treatment  of  Sterility  in  the  Woman. 

By  Dr.  De  Sinety. 

Bacterial  Poisons. 

By  N.  Gamalela,  M.  D. 


Diagnosis    and    Treatment  of   Surgical 
Affections  of  the  Peripheral  Nerves. 
By  F.  Jenner  Hodges,  M.  D. 

Deformities  of  the  Foot. 

By  B.  E.  McKenzie,  M.  D. 

Treatise  on  Diphtheria.. 
By  H.  Bourges,  M.  D. 

Antiseptic  Therapeutics.    Vol.  I. 

Antiseptic  Therapeutics.    Vol.  II. 

By  E.  Trouessart,  M.  D. 

Treatment  of  Typhoid  Fever. 
By  Juhel-Renoy,  M.  D. 


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